DK2478097T3

DK2478097T3 - STABLE LIQUID CLEANING OR CLEANING PRODUCT CONTAINING A PROTEASE

Info

Publication number: DK2478097T3
Application number: DK10760643.6T
Authority: DK
Inventors: Petra Siegert; Astrid Spitz; Timothy OCONNELL; Inken Prüser; Marc Steffen Schiedel; Mueller Dorota Sendor; Thorsten Bastigkeit; Konstantin Benda; Sven Müller; Thomas Eiting; Karl Heinz Maurer; Susanne Wieland
Original assignee: Basf Se
Priority date: 2009-09-16
Filing date: 2010-09-15
Publication date: 2018-02-26
Also published as: EP2478097B1; JP5819302B2; PL2478097T3; JP2013504676A; WO2011032988A1; ES2661317T3; US9163226B2; DE102009029513A1; US20120238005A1; EP2478097A1; CN106995757A; CN102498207A

Description

The invention is in the field of liquid washing and cleaning compositions. The invention relates in particular to the use of defined proteases for provision of proteolytic activity in a liquid washing or cleaning composition, in combination with a phosphonate, and furthermore proposes methods in which such compositions are used.

For washing and cleaning compositions, preferably proteases of the subtilisin type have been used to date. The proteases used in the washing or cleaning compositions known from the prior art originate either originally from microorganisms, for example the genera Bacillus, Streptomyces, Humicola, or Pseudomonas, and/or are produced by biological processes known per se by means of suitable microorganisms, for example by transgenic expression hosts of the genera Bacillus or by filamentous fungi.

Phosphonates are present in modern liquid washing compositions in particular, but also in cleaning compositions. They are used, for example, as complexing agents, for prevention of precipitation or as bleach stabilizer. As complexing agents, they serve, for example, as water softeners. They can ensheath cations such as Ca2 in the solution and hence alter the chemical characteristics of the cation. In the case of calcium, the property of forming water hardness disappears. It is also possible for other cations to be complexed and hence protected from chemical reactions. They can also be involved in the action of corrosion inhibitors or serve as stabilizers for peroxides, especially in bleaches.

The international patent application WO 95/23221 discloses proteases and protease variants of the subtilisin type from Bacillus lentus DSM 5483 which are suitable for use in washing or cleaning compositions. Among these proteases is also one which can have an amino acid substitution R99E, A, S or G. Furthermore, it is disclosed that the cleaning compositions can comprise a phosphonate, in particular a polyphosphonate. The washing compositions can be solid or liquid. However, this document does not reveal a specific washing or cleaning composition liquid which necessarily comprises a phosphonate in combination with such a protease. A disadvantage of protease-containing liquid washing and cleaning compositions from the prior art is that they often do not have satisfactory proteolytic activity, in particular not at low temperatures, for example between 10°C and 50°C, in particular between 10°C and 40°C or between 20°C and 40°C, and the washing or cleaning composition therefore does not exhibit optimal cleaning performance. A further disadvantage of protease-containing liquid washing and cleaning compositions from the prior art is that they are not adequately storage-stable and they accordingly lose a considerable degree of proteolytic activity within just a short time.

The object of the present invention is to overcome the stated disadvantages and to provide liquid washing or cleaning compositions which have an advantageous proteolytic activity, in particular at temperatures as specified above, and which additionally have improved storage stability.

The invention therefore provides for the use of a protease (al) which comprises an amino acid sequence which is at least 80% identical to the amino acid sequence given in SEQ ID No. 1 and which has at position 99 in the count according to SEQ ID No. 1 the amino acid glutamic acid (E) or aspartic acid (D) , for providing a proteolytic activity in a liquid washing or cleaning composition which also comprises a phosphonate in an amount of from 0.01 to 4% by weight.

Surprisingly, it has been found that a liquid washing or cleaning composition which comprises the combination of such a protease with a phosphonate, has advantageous detergency on protease-sensitive soilings and additionally is advantageously storage-stable. In a preferred embodiment of the invention, the compositions are ones which exhibit advantageous detergency with respect to at least one protease-sensitive soiling at temperatures between 10°C and 60°C, preferably also at low temperatures, for example between 10°C and 50°C, between 10°C and 40°C or between 20°C and 40°C. In one embodiment, the composition therefore allows improved removal of at least one, preferably of more, protease-sensitive soilings on textiles and/or hard surfaces, for example dishes. As regards the international patent application WO 95/23221 mentioned in the introduction, the present invention is therefore a particularly advantageous selection which leads to the obtainment of a high-performance and storage-stable liquid cleaning composition or liquid washing composition.

In the context of the invention, detergency is understood as meaning the lightening performance on one or more soilings, in particular laundry soilings or dishware soilings which are sensitive to degradation by the protease. Examples of such laundry soilings are blood-milk/mascara on cotton, full egg/pigment on cotton, chocolate-milk/mascara on cotton, peanut oil pigment/mascara on polyester/cotton, grass on cotton or cocoa on cotton, in particular in such a way as described below. Examples of such dishware soilings are milk, ground meat or egg yolk. In the context of the invention, both the washing or cleaning composition which comprises the protease and/or the washing or cleaning liquor formed by this composition, as well as the protease itself has a particular detergency. The cleaning performance of the hydrolytic enzyme thus contributes to the detergency of the composition or of the washing or cleaning liquor formed by the composition. The detergency is preferably ascertained as described below.

Washing or cleaning liquor is understood as meaning the use solution comprising the washing or cleaning composition which acts on textiles or fabric (washing liquor) or hard surfaces (cleaning liquor) and thus comes into contact with the soilings present on textiles or fabrics or hard surfaces.

Usually, the washing or cleaning liquor is formed when the washing or cleaning operation starts and the washing or cleaning composition is diluted with water for example in a washing machine, dishwasher or in another suitable container.

In one embodiment of the invention, the washing or cleaning composition comprises a protease comprising an amino acid sequence which is at least 80% identical to the amino acid sequence given in SEQ ID No. 1 and which has the amino acid glutamic acid (E) or aspartic acid (D) at position 99 in the count according to SEQ ID No. 1. With increasing preference the amino acid sequence is at least 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% and very particularly preferably 99% identical to the amino acid sequence given in SEQ ID No. 1. SEQ ID No. 1 is the sequence of the ripe (mature) alkaline protease from Bacillus lentus DSM 5483, which is disclosed in the international patent application WO 92/21760 and to the disclosure of which reference is hereby expressly made.

The protease present in a washing or cleaning composition according to the disclosure comprises an amino acid sequence which is at least 80% identical to the amino acid sequence given in SEQ ID No. 1 and which has the amino acid asparagine (N) or glutamine (Q) or the amino acid alanine (A) or glycine (G) or serine (S) at position 99 in the count according to SEQ ID No. 1. With increasing preference the amino acid sequence is at least 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% and very particularly preferably 99% identical to the amino acid sequence given in SEQ ID No. 1. SEQ ID No. 1 is the sequence of the ripe (mature) alkaline protease from Bacillus lentus DSM 5483, which is disclosed in the international patent application WO 92/21760 and to the disclosure of which reference is hereby expressly made.

According to the invention, it has been found that by adding such a protease to a liquid washing or cleaning composition which comprises a phosphonate, a particularly storage-stable liquid cleaning composition is obtained, in particular as regards its remaining proteolytic activity following storage, in particular following a storage time of 1 to 5 weeks, 1 to 4 weeks, 1.5 to 3 weeks and particularly preferably after 2 weeks . A protease present in a washing or cleaning composition has a proteolytic activity, i.e. it is capable of hydrolyzing peptide bonds of a polypeptide or protein. It is therefore an enzyme which catalyzes the hydrolysis of peptide bonds and as a result is able to cleave peptides or proteins, in particular a subtilisin.

In a further embodiment of the invention, the washing or cleaning composition is one wherein the protease also has at least one of the following amino acids in the count according to SEQ ID No. 1: (a) threonine at position 3 (3T), (b) isoleucine at position 4 (41),

(c) alanine, threonine or arginine at position 61 (61A, 61T or 61R), (d) aspartic acid or glutamic acid at position 154 (154D or 154E), (e) proline at position 188 (188P), (f) methionine at position 193 (193M), (g) isoleucine at position 199 (1991), (h) aspartic acid, glutamic acid or glycine at position 211 (211D, 21IE or 211G), (i) combinations of amino acids (a) to (h).

Besides one of the stated amino acids at position 99, the protease therefore has one or more of the aforementioned amino acids at the respective positions. These amino acids can bring about further advantageous properties and/or further enhance existing properties. In particular, the amino acids mentioned above bring about an increase in the proteolytic activity and/or the stability of the protease in a liquid washing or cleaning composition and/or in the wash liquor formed by this washing or cleaning composition. As a result of adding such a protease to a liquid washing or cleaning composition which comprises a phosphonate, a particularly storage-stable liquid washing composition is therefore likewise obtained, in particular as regards its remaining proteolytic activity following storage, in particular following a storage time of 1 to 5 weeks, 1 to 4 weeks, 1.5 to 3 weeks and particularly preferably after 2 weeks.

The amino acid positions here are defined by an alignment of the amino acid sequence of the protease to be used with the amino acid sequence of the protease from Bacillus lentus, as stated in SEQ ID No. 1. Since the protease from Bacillus lentus in the prior art is an important reference molecule for describing proteases and amino acid changes, it is advantageous, in the assignment of the amino acid positions, to refer to the count of the protease from Bacillus lentus (SEQ ID No. 1). Furthermore, the count follows the ripe (mature) protein. This assignment is to be used in particular also if the amino acid sequence of the protease to be used comprises a higher number of amino acid radicals than the protease from Bacillus lentus according to SEQ ID No. 1.

Proceeding from the specified positions in the amino acid sequence of the protease from Bacillus lentus, the amino acid positions in a protease to be used according to the invention are those which are assigned to these very positions in an alignment.

Besides position 99, particularly advantageous positions are accordingly to be assigned to positions 3, 4, 61, 154, 188, 193, 199 and 211 in an alignment with SEQ ID No. 1 and therefore in the count according to SEQ ID No. 1. In the stated positions, the following amino acid radicals are present in the wild type molecule of the protease from Bacillus lentus: S3, V4, G61, S154, A188, V193, V199, and L211. Particular preference is given to the amino acids 3T, 41, 61A, 154D, 154E, 211D, 211G and 211E, if the corresponding positions in a protease to be used according to the invention are not already naturally occupied by one of these preferred amino acids. The substitutions 3T and 41 lead for example, via a stabilization effect on the molecule, to an improvement in the storage stability and the detergency of the protease and therefore to improved detergency of a phosphonate-containing liquid washing or cleaning composition which comprises the protease .

On the basis of the aforementioned amino acids envisaged for the respective position, further sequence deviations from SEQ ID No. 1 may arise, if SEQ ID No. 1 has another amino acid in the respective position. Depending on the number of sequence deviations from SEQ ID No. 1 present, therefore, different maximum identity values arise which can have a protease to SEQ ID No. 1 to be used according to the invention even if it should agree in all other amino acids with SEQ ID No. 1. This material fact should be taken into consideration for every possible combination of the proposed amino acids in the individual case and is furthermore also dependent on the length of the amino acid sequence of the protease. For example, the maximum identity in the case of one, two, three, four, five, six, seven, eight or nine sequence changes is 99.63%, 99.26%, 98.88%, 98.51%, 98.14%, 97.77%, 97.40%, 97.03% and 96.65%, respectively, for an amino acid sequence 269 amino acids in length, or 99.64%, 99.27%, 98.91%, 98.55%, 98.18%, 97.82%, 97.45%, 97.09% or 96.73%, respectively, for an amino acid sequence 275 amino acids in length.

The identity of nucleic acid or amino acid sequences is determined by means of a sequence comparison. Such a comparison takes place by assigning similar sequences in the nucleotide sequences or amino acid sequences to one another. This sequence comparison takes place preferably on the basis of the BLAST algorithm that is usually used and established in the prior art (cf. for example Altschul, S.F., Gish, W., Miller, W., Myers, E.W. & Lipman, D.J. (1990) "Basic local alignment search tool." J. Mol. Biol. 215:403-410, and Altschul, Stephan F.,Thomas L. Madden, Alejandro A. Schaffer, Jinghui Zhang, Hheng Zhang, Webb Miller, and David J. Lipman (1997) : "Gapped BLAST and PSI-BLAST: a new generation of protein database search programs"; Nucleic Acids Res., 25, pp. 3389-3402) and is in principle performed by assigning similar sequences of nucleotides or amino acids in the nucleic acid or amino acid sequences to one another. A tabular assignment of the positions in question is referred to as an alignment. A further algorithm available in the prior art is the FASTA algorithm. Sequence comparisons (alignments), in particular multiple sequence comparisons, are usually generated using computer programs. For example, the Clustal series are often used (cf. for example Chenna et al. (2003) :

Multiple sequence alignment with the Clustal series of programs. Nucleic Acid Research 31, 3497-3500), T-Coffee (cf. for example Notredame et al. (2000) : T-Coffee: A novel method for multiple sequence alignments. J. Mol. Biol. 302, 205-217) or programs which are based on these programs or algorithms. Examples of commonly used programs include Clustal (cf. for example Chenna et al. (2003) : Multiple sequence alignment with the Clustal series of programs. Nucleic Acid Research 31, 3497-3500) or T-Coffee (cf. for example Notredame et al. (2000) : T-Coffee: A novel method for multiple sequence alignments. J. Mol. Biol. 302, 205-217) and also BLAST or FASTA for the database search, or programs based on these programs or algorithms. In the context of the present invention, sequence comparisons and alignments are preferably generated using the computer program Vector NTT® Suite 10.3 (Invitrogen Corporation, 1600 Faraday Avenue, Carlsbad, California, USA) with the preset default parameters.

Such a comparison reveals the similarity of the compared sequences to one another. It is usually quoted in percentage identity, i.e. the fraction of identical nucleotides and amino acid radicals at the same positions or positions corresponding to one another in an alignment. The broader term homology includes in the case of amino acid sequences conserved amino acid substitutions in the consideration, i.e. amino acids with similar chemical activity since these exercise mostly similar chemical activities within the protein. Consequently, the similarity of the compared sequences can therefore also be quoted percent homology or percent similarity. Identity and/or homology data can be given over entire polypeptides or genes or only over individual regions. Homologs or identical regions of different nucleic acid or amino acid sequences are therefore defined by congruities in the sequences. Such regions often have identical functions. They can be small and comprise only a few nucleotides or amino acids. Such small regions often exercise essential functions for the overall activity of the protein. It may therefore make sense to refer sequence congruities only to individual, in some cases small, regions. Unless stated otherwise, however, identity and homology data in the present application refers to the overall length of the nucleic acid or amino acid sequence given in each case.

In a further embodiment of this subject of the invention, the washing or cleaning composition is one wherein the protease comprises an amino acid sequence which is identical to the amino acid sequence given in SEQ ID No. 1 as stated above and which is obtained or is obtainable from a protease according to SEQ ID No. 1 by means of a single or multiple conservative amino acid substitution. The term "conservative amino acid substitution" means the replacement (substitution) of an amino acid radical by another amino acid radical, where this substitution does not lead to a change in the polarity or charge at the position of the substituted amino acid, e.g. the substitution of a nonpolar amino acid radical with a different nonpolar amino acid radical. In the context of the invention, conservative amino acid substitutions comprise for example: G=A=S, I=V=L=M, D=E, N=Q, K=R, Y=F, S=T, G=A=I=V=L=M=Y=F=W=P=S=T.

In a further embodiment of the invention, a washing or cleaning composition is also one wherein its detergency corresponds at least to that of a washing or cleaning composition which includes a protease which comprises an amino acid sequence which corresponds to the amino acid sequence given in SEQ ID No. 2 or SEQ ID No. 3, where the detergency is determined in a washing system which comprises a washing composition in a dose between 4.0 and 8.0 grams per liter of wash liquor and also comprises the protease, where the proteases to be compared are used with identical activity and the detergency toward one or more of the soilings blood-milk/mascara on cotton, full egg/pigment (whole egg/soot) on cotton, peanut oil pigment/mascara on polyester/cotton and grass on cotton, in particular toward one or more of the soilings blood-milk/mascara on cotton: product No. C-05 available from CFT (Center For Testmaterials) B.V. Vlaardingen, The Netherlands - full egg/pigment (whole egg/soot) on cotton: product No. ION available from Firma wfk Testgewebe GmbH; Bruggen-Bracht, Germany, or product C-S-37 available from CFT (Center For Testmaterials) B.V. Vlaardingen, The Netherlands peanut oil pigment/mascara on polyester/cotton: product No. PC-10 available from CFT (Center For Testmaterials) B.V. Vlaardingen, The Netherlands grass on cotton: product No. 164 available from Eidgenossische Material- und Prufanstalt (EMPA)

Testmaterialien AG, St. Gallen, Switzerland, is determined by measuring the degree of whiteness of the washed textiles, the washing operation takes place for at least 30 minutes, optionally 60 minutes, at a temperature of 20°C, and the water has a water hardness between 15.5 and 16.5° (German hardness).

In a further embodiment of the disclosure, a washing or cleaning composition is also one wherein its detergency corresponds at least to that of a washing or cleaning composition which includes a protease which comprises an amino acid sequence which corresponds to the amino acid sequence given in SEQ ID No. 4 or SEQ ID No. 5 or SEQ ID No. 6 or SEQ ID No. 7 or SEQ ID No. 8, where the detergency is determined in a washing system which comprises a washing composition in a dose between 4.0 and 8.0 grams per liter of wash liquor and also comprises the protease, where the proteases to be compared are used with identical activity and the detergency toward one or more of the soilings blood-milk/mascara on cotton, full egg/pigment (whole egg/soot) on cotton, peanut oil pigment/mascara on polyester/cotton and grass on cotton, in particular toward one or more of the soilings blood-milk/mascara on cotton: product No. C-05 available from CFT (Center For Testmaterials) B.V. Vlaardingen, The Netherlands - full egg/pigment (whole egg/soot) on cotton: product No. ION available from Firma wfk Testgewebe GmbH; Bruggen-Bracht,

Germany, or product C-S-37 available from CFT (Center For

Testmaterials) B.V. Vlaardingen, The Netherlands peanut oil pigment/mascara on polyester/cotton: product No. PC-10 available from CFT (Center For Testmaterials) B.V. Vlaardingen, The Netherlands grass on cotton: product No. 164 available from

Eidgenossische Material- und Prufanstalt (EMPA)

The washing composition for the washing system is a liquid washing composition which preferably has the composition specified in table 1 (all data in percent by weight):

Table 1:

The preferred dosage of this liquid washing composition is between 7.0 and 7.5 grams per liter of wash liquor, in particular 7.4 grams per liter of wash liquor. In this respect, the weight of the liquid cleaning composition is determined such that the figures are based on its weight. Washing is preferably carried out in a pH range between pH 8 and pH 10.5, preferably between pH 8 and pH 9.

The degree of whiteness, i.e. the lightening of the soilings, as a measure of the detergency is preferably determined using optical measuring methods, preferably photometrically. An instrument suitable for this purpose is for example Minolta CM508d spectrometer. Usually, the instruments for the measurement are calibrated beforehand with a white standard, preferably a white standard supplied at the same time.

As a result of the activity-identical use of the respective protease, it is ensured that even in the event of a possible divergence of the ratio of active substance to total protein (the values of the specific activity), the respective enzymatic properties, i.e., for example, the detergency on certain soilings, are compared. It is generally the case that a low specific activity can be compensated for by addition of a greater amount of protein. Methods of determining the protease activities are familiar to and are used routinely by the person skilled in the art in the field of enzyme technology. For example, methods of this kind are disclosed in Tenside, volume 7 (1970), p. 125-132. The protease activity is preferably reported in PE (protease units). For example, suitable protease activities are 2.25, 5 or 10 PE (protease units) per mL of wash liquor. However, the protease activity is not equal to zero.

Numerous proteases and in particular Subtilisins are formed as so-called preproteins, i.e. together with a propeptide and a signal peptide, where the function of the signal peptide usually consists in ensuring the elimination of the protease from the cell producing it into the periplasma or the medium surrounding the cell, and the propeptide is usually required for the correct folding of the protease. The signal peptide and the propeptide are usually the N-terminal part of the preprotein. The signal peptide is cleaved from the rest of the protease under natural conditions by means of a signal peptidase. Then, the correct final folding of the protease supported by the propeptide takes place. The protease is then in its active form and cleaves the propeptide itself. After the cleavage of the propeptide, the then ripe (mature) protease, in particular Subtilisin, exerts its catalytic activity without the originally present N-terminal amino acids. For technical applications in general and in particular in the context of the invention, the ripe (mature) proteases, i.e. the enzymes processed after they have been produced, are preferred over the preproteins. The proteases can additionally be modified by the cells producing them after the polypeptide chain has been produced, for example by linkage of sugar molecules, formylations, aminations, etc. Such modifications are post-translational modifications and can, but do not have to, exert an influence on the function of the protease.

Furthermore, the ripe protease can also be shortened at its N-terminal and/or C-terminal end such that a protease shortened compared to SEQ ID No. 1 or SEQ ID No. 2 or SEQ ID No. 3 or SEQ ID No. 4 or SEQ ID No. 5 or SEQ ID No. 6 or SEQ ID No. 7 or SEQ ID No. 8, i.e. a fragment, is present in the washing or cleaning composition according to the invention. All identity data refer in this case to the region in which the particular fragment in an alignment is assigned SEQ ID No. 1. However, in each case, the particular fragment includes the position which is assigned to position 99 in an alignment with SEQ ID No. 1 and has a corresponding amino acid at this position. Advantageously, it also includes one or more of the other above-described positions and has one or more corresponding amino acids there. Furthermore, such a fragment is proteolytically active. A fragment further preferred in this regard comprises an amino acid sequence which, over a length of at least 50 or at least 60, 70, 80, 90, 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, 250, 260, 265, 266, 267 or 268 continuous amino acid positions, agrees with SEQ ID No. 1 or SEQ ID No. 2 or SEQ ID No. 3 or SEQ ID No. 4 or SEQ ID No. 5 or SEQ ID No. 6 or SEQ ID No. 7 or SEQ ID No. 8, taking into consideration the aforementioned amino acids for position 99 and optionally also for positions 3 and/or 4 and/or 61 and/or 154 and/or 188 and/or 193 and/or 199 and/or 211. Particularly preferably, the detergency of a liquid washing or cleaning composition with such a fragment corresponds at least to those of a washing or cleaning composition which includes a protease which comprises an amino acid sequence which corresponds to the amino acid sequence given in SEQ ID No. 2 or SEQ ID No. 3 or SEQ ID No. 4 or SEQ ID No. 5 or SEQ ID No. 6 or SEQ ID No. 7 or SEQ ID No. 8, determined as stated above.

In addition, the proteases present in a composition can be adsorbed onto carrier materials and/or embedded into coating substances in order to protect them from premature inactivation. The protease is then released in the wash liquor, i.e. under application conditions, and can display its proteolytic effect. A further subject matter of the invention provides a liquid washing or cleaning composition which comprises: (a) a protease which is selected from the group consisting of a. protease comprising an amino acid sequence according to SEQ ID No. 2 or SEQ ID No. 3; b. protease which comprises an amino acid sequence modified in at least one position compared to SEQ ID No. 2 or SEQ ID No. 3, where the modification in the count according to SEQ ID No. 1 is selected from the group consisting of: i. threonine at position 3 (3T), ii. isoleucine at position 4 (41), iii. alanine, threonine or arginine at position 61 (61A, 61T or 61R),

iv. aspartic acid or glutamic acid at position 154 (154D or 154E), v. proline at position 188 (188P), vi. methionine at position 193 (193M), vii. isoleucine at position 199 (1991), viii. aspartic acid, glutamic acid or glycine at position 211 (211D, 21IE or 211G), ix. combinations of amino acids (i) to (viii); (b) a phosphonate.

These proteases are very particularly preferably used in a liquid washing or cleaning composition. They are obtained starting from SEQ ID No. 1 by substituting the amino acid arginine at position 99 with the amino acid glutamic acid (E) or aspartic acid (D) in the count according to SEQ ID No. 1. These amino acid sequences are given in the sequence protocol as SEQ ID No. 2 and SEQ ID No. 3. Furthermore, these proteases can have, in addition to the amino acid provided for position 99, one or more of the amino acids explained above in positions 3, 4, 61, 154, 188, 193, 199 and 211 to be assigned in an alignment with SEQ ID No. 1 and therefore in the count according to SEQ ID No. 1. The stated amino acids for these positions also bring about in the case of these proteases further advantageous properties and/or enhance existing properties further. In particular, they bring about an increase in the proteolytic activity and/or the stability of the protease in a liquid washing or cleaning composition or in the wash liquor formed by this washing or cleaning composition. All of the preceding statements - where applicable - apply accordingly to these particularly preferred proteases .

Further disclosed is a liquid washing or cleaning composition comprising: (a) a protease which is selected from the group consisting of a. protease comprising an amino acid sequence according to SEQ ID No. 4 or SEQ ID No. 5 or SEQ ID No. 6 or SEQ ID No. 7 or SEQ ID No. 8; b. protease which comprises an amino acid sequence modified in at least one position compared to SEQ ID No. 4 or SEQ ID No. 5 or SEQ ID No. 6 or SEQ ID No. 7 or SEQ ID No. 8, where the modification in the count according to SEQ ID No. 1 is selected from the group consisting of: i. threonine at position 3 (3T), ii. isoleucine at position 4 (41),

iii. alanine, threonine or arginine at position 61 (61A, 61T or 61R), iv. aspartic acid or glutamic acid at position 154 (154D or 154E), v. proline at position 188 (188P), vi. methionine at position 193 (193M), vii. isoleucine at position 199 (1991), viii. aspartic acid, glutamic acid or glycine at position 211 (211D, 21IE or 211G) , ix. combinations of amino acids (i) to (viii); (b) a phosphonate.

These proteases are very particularly preferably used in a liquid washing or cleaning composition. They are obtained starting from SEQ ID No. 1 by substituting the amino acid arginine at position 99 with the amino acid asparagine (N) or glutamine (Q) or the amino acid alanine (A) or glycine (G) or serine (S) in the count according to SEQ ID No. 1. These amino acid sequences are given in the sequence protocol as SEQ ID No. 4, SEQ ID No. 5, SEQ ID No. 6, SEQ ID No. 7 and SEQ ID No. 8. Furthermore, these proteases can have, in addition to the amino acid provided for position 99, one or more of the amino acids explained above in positions 3, 4, 61, 154, 188,

193, 199 and 211 to be assigned in an alignment with SEQ ID

No. 1 and therefore in the count according to SEQ ID No. 1. The stated amino acids for these positions also bring about in the case of these proteases further advantageous properties and/or enhance existing properties further. In particular, they bring about an increase in the proteolytic activity and/or the stability of the protease in a liquid washing or cleaning composition or in the wash liquor formed by this washing or cleaning composition. All of the preceding statements - where applicable - apply accordingly to these particularly preferred proteases.

Phosphonates are salts and organic compounds, in particular esters, of phosphonic acid. Salts that exist are primary (ΜΉ2Ρ03 or HP (0) (OH) (OM') ) and secondary (NH2HP03 or HP(0)(0M')2) phosphonates, where M' is a monovalent metal. These inorganic phosphonates and are also referred to as primary and secondary phosphites, respectively. Inorganic phosphonates are formed for example by reacting phosphonic acid HP (0) (OH)2, in particular the stable tautomeric form of phosphorous acid, with a (primary) or two (secondary) equivalents of base, for example alkali metal hydroxide.

In the context of the present invention, preference is given to organically P-substituted phosphonates which have a phosphorous-carbon bond (organophosphorous compounds). Their general structure is R1P(0)(OR2)2, where R1 and/or R2 = alkyl, aryl or H, where the alkyl or aryl radicals have further substitutions or can carry further chemical groups. Organically P-substituted phosphonates are formed for example by Michaelis-Arbusov reaction. Many of these phosphonates are soluble in water. Some technically important phosphonates also carry amino group(s) in the type NR-(CH2)x-PO(OH)2 (R = alkyl, aryl or H) . Some of these aminophosphonates have structural similarities with complexing agents such as EDTA, NTA or DTPA and have a similar function.

In the context of the present invention, particularly preferred phosphonates include in particular organophosphonates such as, for example, 1-hydroxyethane-l,1-diphosphonic acid (HEDP), aminotri(methylenephosphonic acid) (ATMP, also referred to as aminotris(methylenephosphonic acid) or nitrolotris(methylenephosphonic acid) (NTMP)), diethylenetriaminepenta(methylene-phosphonic acid) (DTPMP or DETPMP or DTPNT), ethylenediaminetetra(methylenephosphonic acid) (EDTMP, also referred to as ethylenediaminetetra(methylenephosphonic acid) and 2-phosphonobutane-1,2,4-tricarboxylic acid (PBS-AM, also referred to as 2-phosphonobutane-l,2,4-tricarboxylic acid or 3-carboxy-3-phosphonoadipic acid), which are mostly used in the form of their ammonium or alkali metal salts. Particular preference is given to diethylenetriaminepenta(methylenephosphonic acid) sodium, in particular for washing compositions in the context of the invention, and/or 1-hydroxyethane-l,1-diphosphonic acid (HEDP), in particular for dishwashing compositions according to the invention and among these in particular machine dishwashing compositions. Phosphonates of this kind are available for example under the trade names Dequest® 2066 and Dequest® 2010, each from Thermphos).

In a preferred embodiment of the invention, the washing or cleaning composition is one which the phosphonate is present in an amount of 0.01 to 4% by weight. Further preferred amounts of the phosphonate present in the washing or cleaning composition are from 0.01% to 3% by weight, from 0.01% to 2.5% by weight, from 0.02% to 2.4% by weight, from 0.02% to 2% by weight, from 0.03% to 1.5% by weight or from 0.05% to 1% by weight.

In one embodiment of the invention, the protease is present in a washing or cleaning composition preferably in an amount of 1 x 10-8 to 5 percent by weight, based on active protein. With increasing preference, the protease is present in the composition in an amount of 0.001% to 5% by weight, further preferably from 0.01% to 5% by weight, even further preferably from 0.05% to 4% by weight and more preferably from 0.075% to 3.5% by weight. The protein concentration can be determined with the help of known methods, for example the BCA method (bicinchoninic acid; 2,2'-bichinolyl-4,4'-dicarboxylic acid) or the biuret method (A. G. Gornall, C. S. Bardawill and M.M. David, J. Biol. Chem., 177 (1948), pp. 751-766).

In a further embodiment of the invention, the washing or cleaning composition is one wherein it also comprises a component which is selected from i. anionic and/or polyanionic substance, and/or ii. cationic and/or polycationic substance, and/or iii. substance having hydroxyl and/or polyhydroxyl group(s).

It has been found that the addition of such substances further improves the detergency of washing and cleaning compositions, in particular of liquid washing or cleaning compositions, which comprise comprise proteases, in particular those as described above, in particular at relatively low temperatures, in particular between 10°C and 50°C, between 10°C and 40°C, between 10°C and 30°C and/or between 20°C and 40°C.

The substances given above under i. are anionic or polyanionic substances, i.e. these substances carry at least one and preferably two or more negative charges. Preferably, it is a polymer with at least one negatively charged monomer, preferably with two or more negatively charged monomers. According to the invention, this polymer is therefore preferably a negatively charged polymer. Preference is given for example to polymers of organic acids or salts thereof, in particular polyacrylates and/or poly-sugar acids and/or polyacrylate copolymers and/or poly-sugar copolymers. Further preferred compounds in this regard are polyacrylsulfonates or polycarboxylates and their salts, copolymers or salts of the copolymers .

Examples of substances to be used particularly preferably are Acusol 587D (polyacrylsulfonate; Rohm & Haas/Dow Chemical), Acusol 445N (polycarboxylate sodium salt; Rohm & Haas/Dow Chemical), Acusol 590 (polyacrylate copolymer; Rohm & Haas/Dow Chemical), Acusol 916 (polyacrylate sodium salt Rohm & Haas/Dow Chemical), Sokalan CP42 (modified polycarboxylate sodium salt; BASF), Sokalan PA 30CL (polycarboxylate sodium salt; BASF), Dequest P 9000 (polymaleic acid; Thermphos), alginic acid, poly-2-acrylamido-2-methyl-l-propanesulfonic acid, poly-4-styrenesulfonic acid-co-maleic acid sodium salt, polyacrylamido-coacrylic acid sodium salt, poly-methacrylic acid sodium salt, poly-methyl vinyl ether-alt maleic acid or polyvinylsulfonic acid sodium salt.

The substances given under ii. are cationic or polycationic substances, i.e. these substances carry at least one and preferably two or more positive charges. Preferably, it is a polymer with at least one positively charged monomer, preferably with two or more positively charged monomers. According to the invention, this polymer is therefore preferably a positively charged polymer. Examples of compounds preferred in this respect are salts of polyamines, polyethyleneimines and copolymers thereof, salts of polyallylamines, salts of the polydiallyldimethylammonium compounds or poly(acrylamide-co-diallyldimethylammonium compounds .

The substances given under iii. are substances which have at least one hydroxyl and/or polyhydroxyl group and preferably two or more hydroxyl and/or polyhydroxyl groups. In this regard, preference is given for example to polyvinyl alcohols, for example those which are available under the trade name Mowiol (Kremer Pigmente GmbH & Co. KG).

At this point, it is expressly indicated that a specific substance can belong to one or more of the aforementioned groups i. to iii. For example, it may be an anionic polymer which has one or more hydroxyl and/or polyhydroxyl group(s). Such a substance then belongs to groups i. and iii. Similarly, a cationic polymer which has one or more hydroxyl and/or polyhydroxyl group(s) belongs to groups ii. and iii.

In the context of the present invention, it is likewise possible to use derivatives of the substances specified above as belonging to i., ii. or iii. In the context of the present application, a derivative is understood as meaning a substance which, proceeding from one of the aforementioned substances, has been chemically modified, for example by means of converting a side chain or by covalent bonding of another compound to the substance. Such a compound can be for example low molecular compounds such as lipids or mono-, oligo- or polysaccharides or amines or amine compounds. Furthermore, the substance can be glycosylated, hydrolyzed, oxidized, N-methylated, N-formylated, N-acetylated or comprise methyl, formyl, ethyl, acetyl, t-butyl, anisyl, benzyl, trifluoroacetyl, N-hydroxysuccinimides, t-butyloxycarbonyl, benzoyl, 4-methylbenzyl, thioanizyl, thiocresyl, benzyloxymethyl, 4-nitrophenyl, benzyloxycarbonyl, 2-nitrobenzoyl, 2-nitrophenylsulfenyl, 4-toluenesulfonyl, pentafluorophenyl, diphenylmethyl, 2-chlorobenzyloxycarbonyl, 2,4,5-trichlorophenyl, 2-bromobenzyloxycarbonyl, 9-fluorenylmethyloxycarbonyl, triphenylmethyl, 2,2,5,7,8-pentamethylchroman-6-sulfonyl. Likewise, a derivative is to be understood as meaning the covalent or noncovalent bonding of the substance to a macromolecular carrier, just like a noncovalent inclusion into suitable macromolecular cage structures. Couplings with other macromolecular compounds, such as for example polyethylene glycol, can also be undertaken. Further preferred chemical modifications are the modification of one or more of the chemical groups -COOH, -OH, =NH, -NH2 -SH to -COOR, -OR, -NHR, -NR2, -NHR, -NR, -SR; where : R is—CH=CH—R2, -C=C-R2, -C(R2)=CH2, -C(R2)=C(R3), -CH=NR2, - C(R2)=N—R3, a 4-7 carbon ring system with or without substitution, a 4-7 nitrogen heterocycle with or without substitution, or a C2 to Cs chain with 1 to 5 double or triple bonds with substitutions selected from RI, R2, or R3, where - R1 is H, —R, —N02, —CN, halide substituent, — N3, —Cl-8 alkyl, - (CH2) nC02R2, -C2-8 alkenyl-C02R2, -0 (CH2) nC02R2, - C(0)NR2R3, —P(0)(OR2)2, alkyl substituted tetrazol-5-yl, - (CH2) nO (CH2) n aryl, -NR2R3, -(CH2)nOR2, -(CH2)nSR2, - N (R2 ) C (0) R3, —S (02) NR2R3, —N (R2 ) S (02) R3, -(CHR2)n NR2R3, - C(0)R3, (CH2) η N(R3)C(0)R3, -N(R2)CR2R3, substituted or unsubstituted (CH2)n-cycloalkyl, substituted or unsubstituted (CH2)n-phenyl, or cycle; where n is a number greater than 1; - R2 is H, halide substituent, -alkyl, -haloalkyl, —(CH2)n-phenyl, —(CH2) 1-3-biphenyl, - (CH2) 1-4-Ph-N (S02-Cl-2-alkyl) 2, -CO (CHR1) n-ORl, -(CHRl)n-heterocycle, -(CHR1)n-NH-CO-Rl, -(CHR1)n-NH-S02Rl, -(CHR1)n-Ph-N(S02-C1-2- alkyl) 2, —(CHR1)n—C(0)(CHR1)—NHRI, -(CHR1)n-C(S)(CHR1)-NHRI, - (CH2) nO (CH2) nCH3, -CF3, -C2-C5 acyl, -(CHRl)nOH, -(CHR1)nC02Rl, -(CHR1)η-0-alkyl, -(CHR1)n-0-(CH2) η-0-alkyl, -(CHR1)n-S-alkyl, -(CHR1)n-S(0)- alkyl, —(CHR1)n—S (O2)-alkyl, - (CHR1) n—S (02) —NHR3, -(CHR3)n-N3, — (CHR3) nNHR4 , a C2 to C8 alkene chain with 1 to 5 double bonds, a C2 to C8 alkyne chain with 1 to 5 triple bonds, substituted or unsubstituted -(CHR3)n heterocycle, substituted or unsubstituted saturated or unsaturated -(CHR3)n cycloalkyl; where n is a number greater than 1 and R1 and R3 can be identical or different; - R3 is H, —OH, —CN, substituted alkyl, — C2 to C8 alkenyl, substituted or unsubstituted cycloalkyl, —N(R1)R2, saturated or unsaturated C5 to C7 heterocycle or heterobicycle of 4 to 7 carbon atoms, —NR 1, —NR2, —NR1R2 consisting of a saturated or unsaturated heterocycle or a heterobicycle of 4 to 7 carbon atoms; - R4 is H, — (CH2) nOH, -C(0)0R5, -C(0)SR5, — (CH2) nC (0) NR6R7, -0- C (0) —0—R6, an amino acid or a peptide; where n is a number between 0 and 4; - R5 is H; - R6 is -C (R7)-(CH2) n-O-C (0)-R8, - (CH2) n-C (R7)-0-C (0) R8, - (CH2) n-C (R7 )-0-0 (0)-0-R8, or -C (R7)-(CH2) n-O-C (0)-0-R8; where n is a number between 0 and 4; and - R7 and R8 are in each case H, alkyl, substituted alkyl, aryl, substituted aryl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, heterocycle, substituted heterocycle, alkylaryl, substituted alkylaryl, cycloalkyl, substituted cycloalkyl, or CH2C02 alkyl, where R7 and R8 can be identical or different.

According to the invention, it is also possible to use all possible combinations of the substances specified above as belonging to i., ii. or iii. and/or derivatives thereof.

In a preferred embodiment of the invention, the liquid washing or cleaning composition can be used as such or following dilution with water for cleaning textiles and/or hard surfaces. Such a dilution can be easily prepared by diluting a measured-out amount of the composition in a further amount of water in certain weight ratios of composition: water and optionally this dilution is shaken in order to ensure uniform distribution of the composition within the water. Possible weight or volume ratios of the dilutions are from 1:0 composition : water to 1:10 000 or 1:20 000 composition : water, preferably from 1:10 to 1:2000 composition : water.

Liquid washing or cleaning compositions that can be used here are all liquid or flowable presentation forms. In the context of the present application, "flowable" here means compositions which can be poured and can have viscosities up to several 10 000 mPas. The viscosity can be measured using customary standard methods (for example Brookfield viscometer LVT-II at 20 rpm and 20°C, spindle 3) and is preferably in the range from 5 to 10 000 mPas. Preferred compositions have viscosities from 10 to 8000 mPas, with values between 120 and 3000 mPas being particularly preferred. A liquid washing or cleaning composition in the context of the present invention can therefore also be gel-like or paste-like; it can be present as homogeneous solutions or suspensions, as well as for example be sprayable or formulated in other customary presentation forms. The washing compositions include all conceivable types of washing composition, in particular washing compositions for textiles, carpets or natural fibers. They can be provided for manual use and/or also machine use. Washing compositions also include washing auxiliaries which are added to the actual washing composition during manual or machine textile washing in order to achieve a further effect. Cleaners include all compositions, likewise occurring in all specified presentation forms, for the cleaning and/or disinfection of hard surfaces, manual and machine dishwashing washing compositions, carpet cleaners, scouring agents, glass cleaners, WC fragrance flushes, etc. Finally, textile pre- and after treatment compositions are on the one hand those compositions with which the item of laundry is brought into contact before the actual wash, for example for removing stubborn stains, and on the other hand are those which, in a step which follows the actual textile washing, gives the washed item further desirable properties such as a pleasant feel, freedom from creasing or low static charging. The last-mentioned compositions include inter alia the fabric softeners. Disinfectants are for example hand disinfectants, surface disinfectants and instrument disinfectants, which can likewise occur in the specified presentation forms.

In a further preferred embodiment of the invention, the washing or cleaning composition is one which comprises at least one further ingredient, in particular one selected from the group consisting of surfactant, builder, peroxygen compound, bleach activator, alcohol, acid, graying inhibitor, optical brightener, foam inhibitor, water-soluble salt, thickener, volatile alkali and/or base, hydrophilizing agent, and combinations thereof.

Surfactant (s) which can be used are anionic, nonionic, zwitterionic and/or amphoteric surfactants. From an applications point of view, preference is given to mixtures of anionic and nonionic surfactants. The total surfactant content of the liquid washing or cleaning composition is preferably below 60% by weight and particularly preferably below 45% by weight, based on the total liquid washing or cleaning composition .

Suitable nonionic surfactants comprise alkoxylated fatty alcohols, alkoxylated fatty acid alkyl esters, fatty acid amides, alkoxylated fatty acid amides, polyhydroxy fatty acid amides, alkylphenol polyglycol ethers, amine oxides, alkyl polyglucosides and mixtures thereof.

Nonionic surfactants that are used are preferably alkoxylated, advantageously ethoxylated, in particular primary alcohols having preferably 8 to 18 carbon atoms and, on average, 1 to 12 mol of ethylene oxide (EO) per mole of alcohol, in which the alcohol radical can be linear or preferably 2-methyl branched and/or can comprise linear or methyl-branched radicals in a mixture, as are usually present in oxo alcohol radicals. In particular, however, alcohol ethoxylates with linear radicals from alcohols of native origin having 12 to 18 carbon atoms, for example from coconut, palm, tallow fatty or oleyl alcohol, and on average 2 to 8 EO per mole of alcohol are preferred. The preferred ethoxylated alcohols include, for example, Ci2-i4-alcohols with 3 EO, 4 EO or 7 EO, Cg-n-alcohol with 7 EO, Ci3-i5-alcohols with 3 EO, 5 EO, 7 EO or 8 EO, C12-18-alcohols with 3 EO, 5 EO or 7 EO and mixtures of these, such as mixtures of Ci2-i4-alcohol with 3 EO and Ci2-i8-alcohol with 7 EO. The stated degrees of ethoxylation are statistical average values which may be an integer or a fraction for a specific product. Preferred alcohol ethoxylates have a narrowed homolog distribution (narrow range ethoxylates, NRE) . In addition to these nonionic surfactants, fatty alcohols with more than 12 EO can also be used. Examples thereof are tallow fatty alcohol having 14 EO, 25 EO, 30 EO or 40 EO. Nonionic surfactants which comprise EO and PO groups together in the molecule can also be used according to the invention. In addition, also of suitability are a mixture of a (more greatly) branched ethoxylated fatty alcohol and an unbranched ethoxylated fatty alcohol, such as, for example, a mixture of a Ci6-i8-fatty alcohol with 7 EO and 2-propylheptanol with 7 EO. Particularly preferably, the washing composition, cleaner, after-treatment or washing auxiliary comprises a Ci2-i8_fatty alcohol with 7 EO or a C13-15-OXO alcohol with 7 EO as nonionic surfactant.

The content of nonionic surfactants in the washing or cleaning composition is preferably 3 to 40% by weight, preferably 5 to 30% by weight and in particular 7 to 20% by weight, in each case based on the total washing or cleaning composition.

Besides the nonionic surfactants, the washing or cleaning composition can also comprise anionic surfactants. The anionic surfactants used are preferably sulfonates, sulfates, soaps, alkyl phosphates, anionic silicone surfactants and mixtures thereof .

The surfactants of the sulfonate type contemplated here are preferably C9-i3-alkylbenzenesulf onates, olef insulfonates, i.e. mixtures of alkene- and hydroxyalkane sulfonates, and disulfonates, as are obtained, for example from C12-18-monoolefins with terminal or internal double bond by sulfonation with gaseous sulfur trioxide and subsequent alkaline or acidic hydrolysis of the sulfonation products. Also of suitability are Ci2-i8-alkanesulf onates and the esters of a-sulfo fatty acids (ester sulfonates), for example the a-sulfonated methyl esters of the hydrogenated coconut, palm kernel or tallow fatty acids.

Preferred alk(en)yl sulfates are the alkali metal and in particular the sodium salts of the sulfuric acid half-esters of the Ci2-Cis-fatty alcohols, for example of coconut fatty alcohol, tallow fatty alcohol, lauryl, myristyl, cetyl or stearyl alcohol or of the C10-C20-OXO alcohols and those halfesters of secondary alcohols of these chain lengths. From a washing point of view, the Ci2-Ci6_alkyl sulfates and C12-C15-alkyl sulfates, as well as Ci4-Cis-alkyl sulfates are preferred. 2,3-Alkyl sulfates are also suitable anionic surfactants.

Sulfuric acid monoesters of the straight-chain or branched C7-21-alcohols ethoxylated with 1 to 6 mol of ethylene oxide, such as 2-methyl-branched C9-n-alcohols having on average 3.5 mol of ethylene oxide (EO) or Ci2-i8_fatty alcohols with 1 to 4 EO, are also suitable.

Preferred anionic surfactants are also soaps. Of suitability are saturated and unsaturated fatty acid soaps, such as the salts of lauric acid, myristic acid, palmitic acid, stearic acid, (hydrogenated) erucic acid and behenic acid, and also soap mixtures derived in particular from natural fatty acids, for example coconut, palm kernel, olive oil or tallow fatty acid.

The anionic surfactants including the soaps can be present in the form of their sodium, potassium or magnesium or ammonium salts. Preferably, the anionic surfactants are present in the form of their sodium salts. Further preferred counter ions for the anionic surfactants are also the protonated forms of choline, triethylamine or methylethylamine.

The content of anionic surfactants in a washing or cleaning composition can be 1 to 40% by weight, preferably 5 to 30% by weight and very particularly preferably 10 to 25% by weight, in each case based on the total washing or cleaning composition .

Builders which can be present in the washing or cleaning composition are in particular silicates, aluminum silicates (in particular zeolites), carbonates, salts of organic di- and polycarboxylic acids, and mixtures of these substances.

Organic builders which can be present in the washing or cleaning composition are, for example, the polycarboxylic acids that can be used in the form of their sodium salts, where polycarboxylic acids are understood as meaning carboxylic acids which carry more than one acid function. For example, these are citric acid, adipic acid, succinic acid, glutaric acid, malic acid, tartaric acid, maleic acid, fumaric acid, sugar acids, aminocarboxylic acids, nitrilotriacetic acid (NTA), methyl glycine diacetic acid (MGDA) and their modifications, as well as mixtures of these. Preferred salts are the salts of the polycarboxylic acids such as citric acid, adipic acid, succinic acid, glutaric acid, tartaric acid, sugar acids and mixtures of these.

Polymeric polycarboxylates are further suitable as builders, these are for example the alkali metal salts of polyacrylic acid or of polymethacrylic acid, for example those with a relative molecular mass from 600 to 750 000 g / mol.

Suitable polymers are in particular polyacrylates which preferably have a molecular mass of 1000 to 15 000 g/mol. On account of their superior solubility, preference from this group may in turn be given to the short-chain polyacrylates which have molar masses from 1000 to 10 000 g/mol, and particularly preferably from 1000 to 5000 g/mol.

Also of suitability are copolymeric polycarboxylates, in particular those of acrylic acid with methacrylic acid and of acrylic acid or methacrylic acid with maleic acid. To improve the solubility in water, the polymers can also comprise allylsulfonic acids, such as allyloxybenzenesulfonic acid and methallylsulfonic acid, as monomer.

Preferably, however, soluble builders, such as, for example, citric acid, or acryl polymers with a molar mass from 1000 to 5000 g/mol are preferably used in the liquid washing or cleaning compositions.

Organic builder substances of this kind can be present if desired in amounts up to 40% by weight, in particular up to 25% by weight and preferably from 1% by weight to 8% by weight. Amounts close to the specified upper limit are preferably used in pasty or liquid, in particular water-containing, compositions.

Peroxygen compounds suitable for use in compositions according to the invention are in particular organic peracids or peracidic salts of organic acids, such as phthalimido-percaproic acid, perbenzoic acid or salts of diperdodecanedioic acid, hydrogen peroxide and inorganic salts which release hydrogen peroxide under the washing conditions, which include perborate, percarbonate, persilicate and/or persulfate such as Caroat. If a composition comprises peroxygen compounds, these are present in amounts of preferably up to 50% by weight, in particular from 5% by weight to 30% by weight. The addition of small amounts of known bleach stabilizers such as, for example, of phosphonates, borates or metaborates and metasilicates, and also magnesium salts such as magnesium sulfate may be appropriate .

Bleach activators which can be used are compounds which produce, under perhydrolysis conditions, aliphatic peroxocarboxylic acids having preferably 1 to 10 carbon atoms, in particular 2 to 4 carbon atoms, and/or optionally substituted perbenzoic acid. Substances which carry 0- and/or N-acyl groups of the specified number of carbon atoms and/or optionally substituted benzoyl groups are suitable. Preference is given to polyacylated alkylenediamines, in particular tetraacetylethylenediamine (TAED), acylated triazine derivates, in particular 1,5-diacetyl-2,4-dioxohexahydro-1,3,5-triazine (DADHT), acylated glycolurils, in particular tetraacetylglycoluril (TAGU), N-acylimides, in particular N-nonanoylsuccinimide (NOSI), acylated phenolsulfonates, in particular n-nonanoyl- or isononanoyloxybenzenesulfonate (n-or iso-NOBS), carboxylic anhydrides, in particular phthalic anhydride, acylated polyhydric alcohols, in particular triacetin, ethylene glycol diacetate, 2,5-diacetoxy-2,5-dihydrofuran and enol esters, and also acetylated sorbitol and mannitol or their described mixtures (SORMAN), acylated sugar derivatives, in particular pentaacetylglucose (PAG) , penta-acetylfructose, tetraacetylxylose and octaacetyllactose, and also acetylated, optionally N-alkylated glucamine and gluconolactone, and/or N-acylated lactams, for example N-benzoylcaprolactam. The hydrophilically substituted acylacetals and the acyllactams are likewise preferably used. Combinations of conventional bleach activators can also be used. Bleach activators of this type can be present, especially in the case of the presence of aforementioned hydrogen peroxide-producing bleaches, in the customary quantitative range, preferably in amounts of 0.5% by weight to 10% by weight, in particular 1% by weight to 8% by weight, based on the total composition, but are preferably omitted entirely when using percarboxylic acid as the sole bleach.

In addition to the conventional bleach activators, or instead of them, it is also possible for sulfonimines and/or bleachboosting transition metal salts or transition metal complexes to be present as so-called bleach catalysts.

The washing or cleaning compositions are liquid and comprise preferably water as main solvent. In addition, nonaqueous solvents can be added to the washing or cleaning composition. Suitable nonaqueous solvents comprise mono- or polyhydric alcohols, alkanolamines or glycol ethers provided they are miscible in the stated concentration range with water. The solvents are preferably selected from ethanol, n-propanol, i-propanol, butanols, glycol, propanediol, butanediol, glycerol, diglycol, propyl diglycol, butyl diglycol, hexylene glycol, ethylene glycol methyl ether, ethylene glycol ethyl ether, ethylene glycol propyl ether, ethylene glycol mono-n-butyl ether, diethylene glycol methyl ether, diethylene glycol ethyl ether, propylene glycol methyl ether, propylene glycol ethyl ether, propylene glycol propyl ether, dipropylene glycol monomethyl ether, dipropylene glycol monoethyl ether, diisopropylene glycol monomethyl ether, diisopropylene glycol monoethyl ether, methoxytriglycol, ethoxytriglycol, butoxytriglycol, l-butoxyethoxy-2-propanol, 3-methyl-3- methoxybutanol, propylene glycol t-butyl ether, di-n-octyl ether, and mixtures of these solvents. However, it is preferred that the washing or cleaning composition comprises a polyol as nonaqueous solvent. The polyol can comprise in particular glycerol, 1,2-propanediol, 1,3-propanediol, ethylene glycol, diethylene glycol and/or dipropylene glycol. In particular, the washing or cleaning composition preferably comprises a mixture of a polyol and a monohydric alcohol. Nonaqueous solvents can be used in the washing or cleaning composition in amounts between 0.5 and 15% by weight, but preferably below 12% by weight and.

To establish a desired pH that does not result by itself as a result of mixing the other components, the compositions can comprise system- and environment-compatible acids, in particular citric acid, acetic acid, tartaric acid, malic acid, lactic acid, glycolic acid, succinic acid, glutaric acid and/or adipic acid, but also mineral acids, in particular sulfuric acid, or bases, in particular ammonium or alkali metal hydroxides. pH regulators of this kind are present in the compositions in amounts of preferably not more than 20% by weight, in particular from 1.2% by weight to 17% by weight.

Graying inhibitors have the task of keeping the dirt detached from the textile fiber suspended in the liquor. Of suitability for this are water-soluble colloids, mostly organic in nature, for example starch, size, gelatin, salts of ethercarboxylic acids or ethersulfonic acids of starch or of cellulose or salts of acidic sulfuric acid esters of cellulose or of starch. Water-soluble polyamides comprising acidic groups are also suitable for this purpose. Furthermore, it is possible to use starch derivatives other than those mentioned above, for example aldehyde starches. Preference is given to using cellulose ethers, such as carboxymethylcellose (Na salt) , methylcellulose, hydroxyalkylcellulose and mixed ethers, such as methylhydroxyethylcellulose, methylhydroxypropylcellulose, methylcarboxymethylcellulose and mixtures thereof, for example in amounts of 0.1 to 5% by weight, based on the composition.

Textile washing compositions can comprise, as optical brighteners, for example derivatives of diaminostilbenedisulfonic acid or alkali metal salts thereof, although they are preferably free from optical brighteners for use as color washing compositions. Of suitability are, for example, salts of 4,4'-bis(2-anilino-4-morpholino-l,3,5-triazinyl-6-amino)stilbene-2,2'-disulfonic acid or compounds with the same type of structure which carry a diethanolamino group, a methylamino group, an anilino group or a 2-methoxy-ethylamino group instead of the morpholino group. Furthermore, brighteners of the substituted diphenylstyryl type may be present, for example the alkali metal salts of 4,4'-bis (2- sulfostyryl)diphenyl, 4,4'-bis(4-chloro-3-sulfostyryl) diphenyl, or 4-(4-chlorostyryl)-4'-(2-sulfostyryl) diphenyl. Mixtures of the aforementioned optical brighteners can also be used.

Particularly when used in machine processes, it may be advantageous to add foam inhibitors to the compositions. Suitable foam inhibitors are, for example, soaps of natural or synthetic origin which have a high fraction of Ci8_C24_f atty acids. Suitable non-surfactant-like foam inhibitors are, for example, organopolysiloxanes and mixtures thereof with microfine, optionally silanized silica, and also paraffins, waxes, microcrystalline waxes and mixtures thereof with silanized silica or bis-fatty acid alkylenediamides. Mixtures of different foam inhibitors are also advantageously used, for example those of silicones, paraffins or waxes.

Preferably, the foam inhibitors, in particular silicone-and/or paraffin-containing foam inhibitors, are bonded to a granular, water-soluble or water-dispersible carrier substance. In this connection, particular preference is given to mixtures of paraffins and bistearylethylenediamide.

In the context of the invention, a composition can furthermore comprise one or more water-soluble salts, which serve for example for adjusting the viscosity. These may be inorganic and/or organic salts. Inorganic salts that can be used here are preferably selected from the group comprising colorless water-soluble halides, sulfates, sulfites, carbonates, hydrogen carbonates, nitrates, nitrites, phosphates and/or oxides of the alkali metals, of the alkaline earth metals, of aluminum and/or of the transition metals; ammonium salts can also be used. Particular preference is given here to halides and sulfates of the alkali metals; the inorganic salt is therefore preferably selected from the group comprising sodium chloride, potassium chloride, sodium sulfate, potassium sulfate, and mixtures of these. Organic salts that can be used are for example colorless water-soluble alkali metal, alkaline earth metal, ammonium, aluminum and/or transition metal salts of the carboxylic acids. Preferably, the salts are selected from the group comprising formate, acetate, propionate, citrate, malate, tartrate, succinate, malonate, oxalate, lactate, and mixtures thereof. A composition in the context of the invention can further comprise one or more thickeners for the purposes of thickening. The thickener is preferably selected from the group comprising xanthane, guar, carrageenan, agar-agar, gellan, pectin, carob seed flour and mixtures thereof. These compounds are also effective thickeners in the presence of inorganic salts. In a particularly preferred embodiment, the washing or cleaning composition comprises xanthan as thickener since xanthan also effectively thickens in the presence of high salt concentrations and prevents a macroscopic separation of the continuous phase. Additionally, the thickener stabilizes the continuous, low-surfactant phase and prevents macroscopic phase separation.

Alternatively or additionally, (meth)acrylic acid (co)polymers can also be used as thickeners. Suitable acrylic and methacrylic (co)polymers comprise for example the high molecular weight homopolymers of acrylic acid crosslinked with a polyalkylene polyether, in particular an allyl ether of sucrose, pentaerythritol or propylene (INCI name according to "International Dictionary of Cosmetic Ingredients" of "The Cosmetic, Toiletry and Fragrance Association (CTFA)": Carbomer) , which are also referred to as carboxyvinyl polymers. Such polyacrylic acids are available inter alia under the trade names Polygel® and Carbopol®. Furthermore, the following acrylic acid copolymers, for example, are suitable: (i) copolymers of two or more monomers from the group of acrylic acid, methacrylic acid and their simple esters, preferably formed with Ci-4-alkanols (INCI acrylates copolymer), which are available for example under the trade names Aculyn®, Acusol® or Tega® polymer; (ii) crosslinked high molecular weight acrylic acid copolymers, which include for example the copolymers of Cio-30-alkyl acrylates crosslinked with an allyl ether of sucrose or of pentaerythritol with one or more monomers from the group of acrylic acid, methacrylic acid and their simple esters, preferably formed with C1-4-alkanols (INCI Acrylates/Cio-30 Alkyl Acrylate Crosspolymer) and which are available for example under the trade name Carbopol®. Further suitable polymers are (meth)acrylic acid (co)polymers of the Sokalan® type.

It may be preferred that the washing or cleaning composition comprises a (meth)acrylic acid (co)polymer in combination with a further thickener, preferably xanthan. The washing or cleaning composition can comprise 0.05 to 1.5% by weight and preferably 0.1 to 1% by weight, in each case based on the total washing or cleaning composition, of thickener. The amount of thickener used here is dependent on the type of thickener and the desired degree of thickening. A corresponding composition may further comprise volatile alkali. Ammonia and/or alkanolamines which may contain up to 9 carbon atoms in the molecule are used as such.

Alkanolamines used are preferably the ethanolamines, and among these in turn monoethanolamine. The content of ammonia and/or alkanolamine is preferably 0.01% to 2% by weight; particular preference is given to using ammonia. In addition, it is also possible for small amounts of bases to be present. Preferred bases come from the group of the alkali metal and alkaline earth metal hydroxides and carbonates, especially the alkali metal hydroxides, among which potassium hydroxide and in particular sodium hydroxide are particularly preferred. A corresponding composition may also comprise a hydrophilizing agent. In the context of the present invention, this is understood to mean agents for hydrophilization of surfaces. Suitable agents for hydrophilization are especially colloidal silica sols in which the silicon dioxide is preferably in nanoparticulate form. Colloidal nanoparticulate silica sols in the context of this invention are stable dispersions of amorphous particulate silicon dioxide SiC>2 having particle sizes in the range from 1 to 100 nm. Preferably, the particle sizes here are in the range from 3 to 50 nm, more preferably 4 to 40 nm. An example of a silica sol suitable for use in the context of this invention is the silica sol having a particle size of 9 nm available under the Bindzil® 30/360 trade name from Akzo.

Further suitable silica sols are Bindzil® 15/500, 30/220, 40/200 (Akzo), Nyacol® 215, 830, 1430, 2034DI and Nyacol® DP5820, DP5480, DP5540 etc. (Nyacol Products), Levasil® 100/30, 100F/30, 100S/30, 200/30, 200F/30, 300F/30, VP 4038, VP 4055 (H.C. Starck/ Bayer) or else CAB-O-SPERSE® PG 001, PG 002 (aqueous dispersions of CAB-O-SIL®, Cabot), Quartron PL-1, PL-3 (FusoChemical Co.), Kostrosol 0830, 1030, 1430 (Chemiewerk Bad Kostritz) . The silica sols used may also be surface-modified silica that has been treated with sodium aluminate (alumina-modified silica).

In addition, it is also possible to use particular polymers for hydrophilization of surfaces. Suitable hydrophilizing polymers are especially amphoteric polymers, for example copolymers of acrylic or methacrylic acid and MAPTAC, DADMAC or another polymerizable quaternary ammonium compound. In addition, it is also possible to use copolymers with AMPS (2-acrylamido-2-methylpropanesulfonic acid). Polyethersiloxanes, i.e. copolymers of polymethylsiloxanes having ethylene oxide or propylene oxide segments are further suitable polymers. Likewise usable are acrylic polymers, maleic acid copolymers and polyurethanes with PEG (polyethylene glycol) units. Suitable polymers are commercially available, for example, under the Mirapol Surf-S 100, 110, 200, 210, 400, 410, A 300, A 400 (Rhodia), Tegopren 5843 (Goldschmidt), Sokalan CP 9 (BASF) or Polyquart Ampho 149 (Cognis) trade names.

The ingredients of the composition to be chosen, and also the conditions under which it is employed in accordance with the invention, for example temperature, pH, ionic strength, redox ratios or mechanical effects, are typically optimized for the particular field of use.

Liquid or pasty compositions in the form of solutions comprising customary solvents are generally prepared by simple mixing of the ingredients, which can be added without dilution or as a solution to an automatic mixer.

Washing or cleaning compositions in the context of the invention can exclusively comprise a protease as described. Alternatively, they can also comprise further hydrolytic enzymes or other enzymes in a concentration expedient for the efficacy of the composition. In the context of the invention, compositions can consequently comprise one or more further enzymes, it being possible in principle to use all enzymes established in the prior art for these purposes. Further enzymes that can preferably be used are all enzymes which, in the composition according to the invention, can develop a catalytic activity, in particular a in particular a protease, amylase, cellulase, hemicellulase, mannanase, tannase, xylanase, xanthanase, β-glucosidase, carrageenase, perhydrolase, oxidase, oxidoreductase or a lipase, as well as preferably mixtures thereof. These enzymes are in principle of natural origin; proceeding from the natural molecules, improved variants are available for use in washing and cleaning compositions, which are correspondingly used with preference .

Compositions in the context of the invention comprise enzymes, preferably in total amounts of 1 x 10-8 to 5% by weight, based on active protein. The enzymes are preferably present in compositions at from 0.001% to 5% by weight, further preferably from 0.01% to 5% by weight, even further preferably from 0.05% to 4% by weight and particularly preferably from 0.075% to 3.5% by weight, where each enzyme present may be in the quantitative ratios specified. The enzymes may be adsorbed onto carrier materials and/or embedded into coating substances in order to protect them from premature inactivation. The protein concentration can be determined with the help of known methods, for example the BCA method (bicinchoninic acid; 2,2'-bichinolyl-4,4'-dicarboxylic acid) or the Biuret method (A. G. Gornall, C. S. Bardawill and M.M. David, J. Biol. Chem., 177 (1948), pp. 751-766) .

Particularly preferably, the enzymes exhibit synergistic effects with respect to their action toward certain soilings or stains, i.e. the enzymes present in the product composition are mutually supported in their cleaning performance. Very particularly preferably, such a synergism is present between the protease present according to the invention and a further enzyme of a composition, including in particular between the specified protease and an amylase and/or a mannase and/or a lipase .

Synergistic effects can arise not only between different enzymes, but also between one or more enzymes and further ingredients of the composition.

Among the proteases, those of the subtilisin type are preferred. Examples thereof are the subtilisins BPN' and Carlsberg, the protease PB92, the subtilisins 147 and 309, the alkaline protease from Bacillus lentus, subtilisin DY and the enzymes to be assigned to the subtilases but no longer to the subtilisins in the narrower sense, these being thermitase, proteinase K and the proteases TW3 and TW7. Subtilisin Carlsberg is available in a further developed form under the trade name Alcalase® from Novozymes A/S, Bagsværd, Denmark.

The subtilisins 147 and 309 are sold by Novozymes under the trade names Esperase®, or Savinase®. Derived from the proteases from Bacillus lentus DSM 5483 are the protease variants listed under the name BLAP®. Further usable proteases are, for example, the enzymes available under the trade names Durazym®, Relase®, Everlase®, Nafizym®, Natalase®, Kannase® and Ovozyme® from Novozymes, those available under the trade names Purafect®, Purafect® OxP, Purafect® Prime, Excellase® and Properase® from Genencor, that available under the trade name Protosol® from Advanced Biochemicals Ltd., Thane, India, that available under the trade name Wuxi® from Wuxi Snyder Bioproducts Ltd., China, those available under the trade names Proleather® and Protease P® from Amano Pharmaceuticals Ltd., Nagoya, Japan, and that available under the name Proteinase K-16 from Kao Corp., Tokyo, Japan. Particular preference is also given to using the proteases from Bacillus gibsonii and Bacillus pumilus, which are disclosed in the international patent applications WO 2008/086916 and WO 2007/131656.

Examples of amylases that can be formulated according to the invention are the α-amylases from Bacillus licheniformis, from B. amyloliquefaciens or from B. stearothermophilus, and their further developments improved for use in washing or cleaning compositions. The enzyme from B. licheniformis is available from Novozymes under the name Termamyl® and from Genencor under the name Purastar®ST. Further-development products of this α-amylase are available from Novozymes under the trade names Duramyl® and Termamyl®ultra, from Genencor under the name Purastar®OxAm and from Daiwa Seiko Inc., Tokyo, Japan, as Keistase®. The α-amylase from B. amyloliquefaciens is sold by Novozymes under the name BAN®, and derived variants from the α-amylase from B. stearothermophilus under the names BSG® and Novamyl®, likewise from Novozymes.

Furthermore, the α-amylase from Bacillus sp. A 7-7 (DSM 12368) and the cyclodextrin-glucanotransferase (CGTase) from B. agaradherens (DSM 9948) are to be emphasized for this purpose. In addition, the amylolytic enzymes which belong to the sequence range of α-amylases which is defined in the international patent application WO 03/002711 A2 and those which are described in the application WO 03/054177 A2 are also usable. Fusion products of the stated molecules can likewise be used.

Moreover, the further developments of the α-amylase from

Aspergillus niger and A. oryzae available under the trade names Fungamyl® from Novozymes are suitable. Further commercial products that can be used are, for example, the Amylase-LT® and Stainzyme® or Stainzyme ultra® or Stainzyme plus®, the latter likewise from Novozymes. Variants of these enzymes obtainable by point mutations can also be used according to the invention.

Examples of lipases or cutinases which can be formulated according to the invention that are present in particular on account of their triglyceride-cleaving activities, but also in order to generate peracids in situ from suitable precursors are the lipases originally obtainable, or further developed, from Humicola lanuginosa (Thermomyces lanuginosus), in particular those with the amino acid exchange D96L. They are sold for example by Novozymes under the trade names Lipolase®, Lipolase®Ultra, LipoPrime®, Lipozyme® and Lipex®.

Furthermore, the cutinases which have originally been isolated from Fusarium solani pisi and Humicola insolens for example can be used. Lipases that are likewise usable are available from Amano under the Lipase CE®, Lipase P®, Lipase B®, or Lipase CES®, Lipase AKG®, Bacillis sp. Lipase®, Lipase AP®, Lipase M-AP® and Lipase AML® names. From Genencor, for example, the lipases or cutinases can be used whose starting enzymes have originally been isolated from Pseudomonas mendocina and Fusarium solanii. Further important commercial products include the preparations Ml Lipase® and Lipomax® originally sold by Gist-Brocades and the enzymes sold by Meito Sangyo KK, Japan, under the names Lipase MY-30®, Lipase OF® and Lipase PL®, also the product Lumafast® from Genencor.

Washing or cleaning compositions in the context of the invention may further comprise cellulases, according to the purpose in the form of pure enzymes, of enzyme preparations or in the form of mixtures in which the individual components are advantageously complementary in terms of their different performance aspects. These performance aspects in particular include contributions to the primary washing performance, to the secondary washing performance of the composition (antiredeposition effect or graying inhibition) and hand (fabric action), up to and including the exertion of a "stonewashed" effect. A usable fungal endoglucanase (EG)-rich cellulose preparation or its further developments is supplied by Novozymes under the trade name Celluzyme®. The products Endolase® and Carezyme® likewise available from Novozymes are based on the 50 kD-EG, or the 43 kD-EG from H. insolens DSM 1800. Further usable commercial products from this company are Cellusoft®, Renozyme® and Celluclean®. Also usable are for example the 20 kD-EG from Melanocarpus which are available from AB Enzymes, Finland, under the trade names Ecostone® and Biotouch®. Further commercial products from AB Enzymes are Econase® and Ecopulp®. Further suitable cellulases are from Bacillus sp. CBS 670.93 and CBS 669.93, where the CBS 670.93 from Bacillus sp. is available from Genencor under the trade name Puradax®. Further commercial products from Genencor are "Genencor detergent cellulase L" and IndiAge®Neutra.

Furthermore, further enzymes can be used in particular for removing certain problem stains; these are summarized under the term hemicellulases. These include for example mannanases, xanthan lyases, pectin lyases ( = pectinases), pectin esterases, pectate lyases, xyloglucanases ( = xylanases), pullulanases and β-glucanases. Enzymes suitable in this regard are available, for example, under the Gamanase® and Pektinex AR® names from Novozymes, under the Rohapec® B1L name from AB Enzymes and under the Pyrolase® name from Diversa Corp., San Diego, CA, USA. The β-glucanase obtained from Bacillus subtilis is available under the Cereflo® name from Novozymes. Hemicellulases that are particularly preferred according to the invention are mannanases, which are sold for example under the trade names Mannaway® by Novozymes or Purabrite® by Genencor.

To increase the bleaching effect, compositions in the context of the invention can also comprise oxidoreductases, for example oxidases, oxygenases, catalases (which react at low H2O2 concentrations as peroxidase), peroxidases, such as haloperoxidases, chiaroperoxidases, bromoperoxidases, lignoperoxidases, glucose peroxidases or manganese peroxidases, dioxygenases or laccases (phenol oxidases, polyphenol oxidases). Suitable commercial products include Denilite® 1 and 2 from Novozymes. As advantageously usable example systems of an enzymatic perhydrolysis, reference is made to the applications WO 98/45398 Al, WO 2005/056782 A2 and WO 2004/058961 Al. A combined enzymatic bleaching system comprising an oxidase and a perhydrolase is described in the application WO 2005/124012. Advantageously, organic, particularly preferably aromatic, compounds which interact with the enzymes are preferably additionally added in order to boost the activity of the oxidoreductases in question (enhancers) or in order to ensure the flow of electrons in the case of very different redox potentials between the oxidizing enzymes and the stains (mediators).

The enzymes used according to the invention either come originally from microorganisms, for instance from the genera Bacillus, Streptomyces, Humicola or Pseudomonas, and/or are produced via suitable microorganisms by biotechnology methods known per se, for instance via transgenic expression hosts from the genera Bacillus or via filamentous fungi. The enzymes in question are favorably purified by means of methods that have become established per se, for example by means of precipitation, sedimentation, concentration, filtration of the liquid phases, microfiltration, ultrafiltration, action of chemicals, deodorization or suitable combinations of these steps. In addition, the enzymes can be formulated together with accompanying substances, for example from the fermentation, or with stabilizers. A particular subject of the invention is the use of a washing or cleaning composition according to the invention for removing soilings, in particular protease-sensitive soilings, on textiles or hard surfaces, i.e. for the cleaning of textiles or of hard surfaces. This is because compositions can, especially on account of the combination of protease and phosphonate present, be advantageously used in order to remove corresponding impurities from textiles or from hard surfaces. Embodiments of this subject of the invention are for example hand washing, the manual removal of stains from textiles or from hard surfaces or the use in conjunction with a machine process .

All material facts, subjects and embodiments which are described for washing or cleaning compositions can also be applied to this subject of the invention. Consequently, at this point, reference is made expressly to the disclosure at the corresponding point with the indication that this disclosure also applies for the above use according to the invention . A further subject of the invention is a method of cleaning textiles or hard surfaces, in which a washing or cleaning composition is used at least in one of the method steps. The method for cleaning textiles or hard surfaces is accordingly one in which, in at least one method step, a washing or cleaning composition is used.

These include both manual and machine methods, preference being given to machine methods on account of their more precise controllability as regards the amounts used and contact times for example.

Methods for cleaning textiles are generally characterized in that in a plurality of method steps, various cleaning-active substances are applied to the ware to be cleaned and, after the contact time, are washed off, or that the ware to be cleaned is treated in some other way with a washing composition or a solution or dilution of this composition. The same is true for methods for cleaning all materials other than textiles, in particular hard surfaces. All conceivable washing or cleaning methods can be enriched in at least one of the method steps by use of a washing or cleaning composition and are then embodiments of the present invention.

All material facts, subjects and embodiments which are described for washing or cleaning compositions are also applicable to this subject of the invention. Consequently, at this point, reference is made expressly to the disclosure at the corresponding point with the indication that this disclosure also applies to the above methods according to the invention .

In a preferred embodiment, the method is one wherein the phosphonate is present in the wash liquor in a concentration of from 0.00075 to 0.05% by weight and/or that the protease is present in the wash liquor in a concentration of from 0.0005 to 0.03% by weight. Further preferred concentrations of the phosphonate present in the wash liquor are from 0.00075 to 0.01125%, from 0.001 to 0.035% by weight, from 0.002 to 0.01125% by weight or from 0.00375 to 0.0075% by weight. Further preferred concentrations of the protease present in the wash liquor are from 0.00075 to 0.03% by weight or from 0.00077 to 0.028% by weight.

In a further preferred embodiment, the method is carried out at a temperature between 10°C and 60°C, between 10°C and 55°C, between 10°C and 50°C, between 10°C and 40°C or between 20°C and 40°C.

According to the above statements, proteases used in compositions can advantageously be used in washing and cleaning compositions in the context of the invention, as well as methods according to the invention, in particular washing and cleaning methods. They can thus advantageously be used in order to provide a proteolytic activity in corresponding compositions .

The invention therefore further provides the use of a protease (al) which comprises an amino acid sequence which is at least 80% identical to the amino acid sequence given in SEQ ID No. 1 and which has the amino acid glutamic acid (E) or aspartic acid (D) at position 99 in the count according to SEQ ID No. 1, for providing a proteolytic activity in a liquid washing or cleaning composition which further comprises a phosphonate in an amount of 0.01 to 4% by weight.

The disclosure also encompasses the use of a protease (a2) comprising an amino acid sequence having at least 80% identity to the amino acid sequence specified in SEQ ID NO. 1 and having, at position 99 in the count according to SEQ ID NO. 1, the amino acid asparagine (N) or glutamine (Q), or (a3) comprising an amino acid sequence having at least 80% identity to the amino acid sequence specified in SEQ ID NO. 1 and having, at position 99 in the count according to SEQ ID NO. 1, the amino acid alanine (A) or glycine (G) or serine (S), for provision of proteolytic activity in a liquid washing or cleaning composition further comprising a phosphonate.

In a further embodiment, this use is one wherein the protease also has at least one of the following amino acids in the count according to SEQ ID No. 1: (a) threonine at position 3 (3T), (b) isoleucine at position 4 (41), (c) alanine, threonine or arginine at position 61 (61A, 61T or 61R),

(d) aspartic acid or glutamic acid at position 154 (154D or 154E), (e) proline at position 188 (188P), (f) methionine at position 193 (193M), (g) isoleucine at position 199 (1991), (h) aspartic acid, glutamic acid or glycine at position 211 (211D, 21IE or 211G), (i) combinations of amino acids (a) to (h).

The invention further provides the use of a protease which is selected from the group consisting of a. protease comprising an amino acid sequence according to SEQ ID No. 2 or SEQ ID No. 3; b. protease which comprises an amino acid sequence modified in at least one position compared to SEQ ID No. 2 or SEQ ID No. 3, where the modification in the count according to SEQ ID No. 1 is selected from the group consisting of: i. threonine at position 3 (3T), ii. isoleucine at position 4 (41), iii. alanine, threonine or arginine at position 61 (61A, 61T or 61R),

iv. aspartic acid or glutamic acid at position 154 (154D or 154E), v. proline at position 188 (188P), vi. methionine at position 193 (193M), vii. isoleucine at position 199 (1991), viii. aspartic acid, glutamic acid or glycine at position 211 (21ID, 21IE or 211G), ix. combinations of amino acids (i) to (viii); for providing a proteolytic activity in a liquid washing or cleaning composition which further comprises a phosphonate in an amount of 0.01 to 4% by weight.

The disclosure further provides the use of a protease which is selected from the group consisting of a. protease comprising an amino acid sequence according to SEQ ID No. 4 or SEQ ID No. 5 or SEQ ID No. 6 or SEQ ID No. 7 or SEQ ID No. 8; b. protease which comprises an amino acid sequence modified in at least one position compared to SEQ ID No. 4 or SEQ ID No. 5 or SEQ ID No. 6 or SEQ ID No. 7 or SEQ ID No. 8, where the modification in the count according to SEQ ID No. 1 is selected from the group consisting of: i. threonine at position 3 (3T), ii. isoleucine at position 4 (41), iii. alanine, threonine or arginine at position 61 (61A, 61T or 61R),

iv. aspartic acid or glutamic acid at position 154 (154D or 154E), v. proline at position 188 (188P), vi. methionine at position 193 (193M), vii. isoleucine at position 199 (1991), viii. aspartic acid, glutamic acid or glycine at position 211 (21ID, 21IE or 211G), ix. combinations of amino acids (i) to (viii); for providing a proteolytic activity in a liquid washing or cleaning composition which further comprises a phosphonate.

All material effects, subjects and embodiments which are described for washing and cleaning compositions are also applicable to said uses. Consequently, at this point, reference is made expressly to the disclosure at the corresponding point with the indication that this disclosure also applies for the above uses according to the invention.

Examples

All molecular biology operations follow standard methods as specified, for example, in the Handbook by Fritsch, Sambrook and Maniatis "Molecular cloning: a laboratory manual", Cold Spring Harbour Laboratory Press, New York, 1989, or comparable standard works. Enzymes and kits were used according to the respective manufacturer's instructions.

Example 1

Determination of the cleaning performance of a liquid washing composition

For this example, soiled textiles were used in a standardized manner, which had been sourced from EMPA Testmaterialien AG (St. Gallen, Switzerland), wfk Testgewebe GmbH (Bruggen-Bracht, Germany) or the Center For Testmaterials (CFT, Vlaardingen, the Netherlands) . The following stains and textiles were used: A: grass on cotton: product No. 164 from the Eidgenossische Material- und Prufanstalt (EMPA) Testmaterialien AG, St. Gallen, Switzerland; B: peanut oil pigment/indian ink on polyester/cotton: product No. PC-10 from CFT (Center For Testmaterials) B.V. Vlaardingen, the Netherlands; C: whole egg/pigment (whole egg/soot) on cotton: product No. ION from wfk Testgewebe GmbH; Bruggen-Bracht, Germany; D: blood-milk/indian ink on cotton: product No. C-05 obtainable from CFT (Center For Testmaterials) B.V. Vlaardingen, the Netherlands

This test material was used to test various washing compositions for their cleaning performance. For this purpose, the batches were washed at temperatures of 40°C or 20°C for 60 minutes. The dosage was 7.4 grams of the washing composition per liter of wash liquor. Washing was effected with mains water having a water hardness of about 16° German hardness. The washing composition used was a washing composition base formulation as specified above in table 1.

For the various test series, this washing composition base formulation was admixed with the following proteases with equal activity (final concentration 5 PE/ml): protease comprising an amino acid sequence according to SEQ ID NO. 2 (batch 1), performance-improved variant F49 of the protease from Bacillus lentus according to WO 95/23221 (batch 2) and the protease disclosed in figure 2 or SEQ ID NO. 3 of international published specification WO 03/057713 (batch 3).

After the wash, the whiteness level of the washed textiles was measured. The measurement was made on a Minolta CM508d spectrometer (D65 illuminant, 10°) . The instrument had been calibrated beforehand with a supplied white standard. The results obtained are the differential reflectances between a washing operation with a washing composition comprising a protease and a control washing operation conducted in parallel with a washing composition lacking protease. The results are compiled in table 2 below and permit a direct conclusion as to the contribution of the enzymes present in each case to the cleaning performance of the composition used.

Table 2: Wash results with a liquid washing composition at 4 0 ° C or 2 0 ° C

Stain Batch 1 Batch 2 Batch 3

__4 0 ° C 2 0 ° C 4 0 ° C 2 0 ° C 40°C 20°C _A__1^4__1^3__lj_4__0_1_4__1^0__0.7 _B__5^2__4^9__4^0__2^9__4^9__4.2 _C__3^0__3^5__4^9__2^3__2Λ__2.4 D I 16.7 I 12.6 I 13.6 |ll.4 |l5.4 |ll.l

It becomes clear that a washing composition in the context of the invention exhibits very good cleaning performance and, on most stains, even a better cleaning performance compared to the washing compositions of batches 2 and 3.

Example 2

Determination of the cleaning performance of a liquid machine dishwashing composition

Vessels having hard smooth surfaces were respectively stained in a standardized manner and washed with a standard commercial domestic machine dishwasher at 40°C and 50°C. In each wash cycle, 30 ml of a standard commercial machine dishwashing composition in liquid or gel form was used, which contained 2.4% by weight of phosphonate (HEDP). Washing was effected with mains water having a water hardness of about 21° German hardness. The protease used was Savinase ultra 16L (Novozymes, batch 1) and an inventive protease comprising an amino acid sequence according to SEQ ID NO. 2 (batch 2) . The proteases were used in equal weights based on enzyme protein (0.68 g of Savinase ultra 16L or the same amount in terms of weight of enzyme protein of the inventive protease per 30 mL of cleaning composition).

After the wash, the removal of the stains was determined either gravimetrically (egg yolk) or visually (further stains according to table 3) . For the gravimetric determination, the difference between the weight of the soiled vessel and the washed vessel was determined (egg yolk released) and this was based on the amount of egg yolk originally applied according to the following formula: % cleaning performance = (mg of egg yolk released / mg of egg yolk applied) x 100. In the visual determination, the soil removal was assigned visually to a percentage scale by persons having experience in this regard. The results are compiled in table 3 below and permit a direct conclusion as to the contribution of the enzyme present in each case to the cleaning performance of the composition used.

Table 3: Cleaning performance of an inventive liquid machine dishwashing composition at 40°C or 50°C

Stain 14 0 ° C i50°C | jBatch 1 ijBatch 2 jBatch 1 ijBatch 2 \

Milk 17 4 17 6 j74 ]7 4 |

Minced |99 1100 i 94 i|98 \ meat \ \ \ \ \

Egg yolk |37 14 7 :4 7 i|65 \

It becomes clear that a liquid machine dishwashing composition in the context of the invention exhibits very good cleaning performance and, particularly on the egg yolk stain, even distinctly better cleaning performance compared to the dishwashing composition according to batch 1.

Example 3

Determination of the storage stability of a liquid washing composition

The washing compositions according to batch 1 and 2 from

Example 1 were tested with regard to their storage stability. For this purpose, the washing compositions were stored at a temperature of 30°C for the period of time specified in each case, and the respective proteolytic residual activity was determined via the release of the para-nitroaniline (pNA) chromophore from the substrate. The substrate is suc-L-Ala-L-Ala-L-Pro-L-Phe-p-nitroanilide (suc-AAPF-pNA). The protease cleaves the substrate and releases pNA. The release of the pNA causes an increase in the absorbance at 410 nm, the course of which over time is a measure of the enzymatic activity (cf. Del Mar et al., 1979). The measurement was made at a temperature of 25°C, at pH 8.6 and at a wavelength of 410 nm. The measurement time was 5 min. with a measurement interval of 20 s to 60 s. The residual activities obtained are reported in table 4 below.

Table 4: Determination of the proteolytic residual activity after storage

Washing composition\Start ijWeek 1 ijWeek 2 j according to \ \ \ \

Batch 1 ] 10 0 % ] 71% ]7 7 % |

Batch 2 1100% ]47% ]37% j

It becomes clear that a washing composition in the context of the invention has a distinct improvement in storage stability compared to a washing composition according to batch 2. The washing composition in the context of the invention has an increase in proteolytic activity by 151% after one week or by 211% after two weeks.

SEQUENCE LISTING

<110> Henkel AG & Co. KGaA <120> Storage-stable liquid washing or cleaning composition comprising proteases

<130> HO8707 PCT <150> DE 102009029513 <151> 2009-09-16 <160> 8 <170> Patentln version 3.3 <210> 1 <211> 269

<212> PRT <213> Bacillus lentus <4 0 0> 1

Ala Gin Ser Val Pro Trp Gly Ile Ser Arg Val Gin Ala Pro Ala Ala IS 10 IS

Hia Asn Arg Gly Leu Thr Gly Ser Gly Val Lys Val Ala Val Leu Asp 20 25 30

Thr Gly Ile Ser Thr His Pro Asp Leu Asn Ile Arg Gly Gly Ala Ser 35 40 45

Pha Val Pro Gly Glu Pro Ser Thr Gin Asp Gly Asn Gly His Gly Thr SO 55 60

His Val Ala Gly Thr He Ala Ala Leu Asn Asn Ser lie Gly Val Leu 65 70 75 80

Gly Val Ala Pro Ser Ala Glu Leu Tyr Ala Val Lys Val Leu Gly Ala 85 90 95

Asp Gly Arg Gly Ala He Ser Ser He Ala Gin Gly Leu Glu Trp Ala 100 105 110

Gly Asn Asn Gly Met His Val Ala Asn Leu Ser Leu Gly Ser Pro Ser 115 120 125

Pro Ser Ala Thr Leu Glu Gin Ala Val Asn Ser Ala Thr Ser Arg Gly 130 135 140

Val Leu Val Val Ala Ala Ser Gly Asn Ser Gly Ala Ser Ser He Ser 145 150 155 160

Tyr Pro Ala Arg Tyr Ala Asn Ala Met Ala Val Gly Ala Thr Asp Gin 165 170 175 Åsn Asn Asn Arg Ala Ser Phe Ser Gin Tyr Gly Ala Gly Le« Asp Ile 180 185 190

Val Ala Pro Gly Val Asn Val Gin Ser Thr Tyr Pro Gly Ser Thr Tyr 195 200 205

Ala Ser Len Asn Gly Thr Ser Met Ala Thr Pro His Val Ala Gly Ala 210 215 220

Ala Ala Len Val Lys Gin Lys Asn Pro Ser Trp Ser Asn Val Gin Ile 225 230 235 240

Arg Asn His Len Lys Asn Thr Ala Thr Ser Leu Gly Ser Thr Asn Len 245 250 255

Tyr Gly Ser Gly Len Val Asn Ala Gin Ala Ala Thr Arg 260 265 <210> 2 <211> 269

<212> PRT <213> Bacillus lentus <400> 2

Ala Gin Ser Val Pro Trp Gly Ile Ser Arg Val Gin Ala Pro Ala Ala 1 5 10 15

His Asn Arg Gly Len Thr Gly Ser Gly Val Lys Val Ala Val Len Asp 20 25 30

Thr Gly Ile Ser Thr His Pro Asp Leu Asn Ile Arg Gly Gly Ala Ser 35 40 45

Phe Val Pro Gly Glu Pro Ser Thr Gin Asp Gly Asn Gly His Gly Thr 50 55 60

His Val Ala Gly Thr Ile Ala Ala Len Asn Asn Ser Ile Gly Val Len 65 70 75 80

Gly Val Ala Pro Ser Ala Gin Leu Tyr Ala Val Lys Val Leu Gly Ala 85 90 95

Asp Gly Glu Gly Ala Ile Ser Ser Ile Ala Gin Gly Len Gin Trp Ala 100 105 110

Gly Asn Asn Gly Met His Val Ala Asn Leu Ser Leu Gly Ser Pro Ser 115 120 125

Pro Ser Ala Thr Leu Glu Gin Åla Val Asa Sar Ala Thr Ser Arg Gly 130 135 140

Val leu Våi Val Ala Ala Ser Gly Åsa Ser Gly Ala Sér Sar 11a Ser

145 150 155 ISO lyr Pro Ala Arg Tyr Ala Asa Ala Mat Ala Val Gly Ala Thr Asp Gin 165 170 175

Asn Asn Asn Arg Ala Ser Phe Ser Gin Tyr Gly Ala Gly Leu Asp lie ISO 185 ISO

Val Ala Pro Gly Val Asn Val Gin Ser Thr Tyr Pro Gly Ser Thr Tyr 195 200 205

Ala Ser leu Asti Gly Thr Ser Mat Ala Thr Pro His Val Ala Gly Ala 210 215 220

Ala Ala leu val Lys Gin I.ys Asn Pro Ser Trp Ser Ash Val Gin lie 225 230 235 240

Arg Asn His Leu Lys Asn Thr Ala Thr Ser Leu Gly Ser Thr Asn Leu 245 250 255

Tyr Gly Ser Gly Leu Val Asn Ala Glu Ala Ala Thr Arg 260 265 <210> 3 <211> 269

<212> PRT <213> Bacillus lentus <400> 3

Ala Gin Ser Val Pro Trp Gly lie Ser Arg Val Gin Ala Pro Ala Ala 1 5 10 15

His Asn Arg Gly Leu Thr Gly Ser Gly Val Lys Val Ala Val Leu Asp 20 25 30

Thr Gly lie Ser Thr His Pro Asp Leu Asn lie Arg Gly Gly Ala Ser 35 40 45

Phe Val Pro Gly Glu Pro Ser Thr Gin Asp Gly Asn Gly His Gly Thr 50 55 60

His val Ala Gly Thr lie Ala Ala Leu Asn Asn Ser lie Gly Val Leu 65 70 75 80

Gly Val Ala Pro Ser Ala Glu Leu Tyr Ala Val Lys Val Leu Gly Ala 35 90 95

Asp Gly Asp Gly Ala Ile Ser Ser lie Ala Gin Gly Leu Glu Trp Ala 100 105 110

Gly Asn Asn Gly Met His Val Ala Asn Leu Ser Leu Gly Ser Fro Ser 115 120 125

Pro Ser Ala Thr Leu Glu Gin Ala Val Asn Ser Ala Thr Ser Arg Gly 130 135 140

Val Leu Val Val Ala Ala Ser Gly Asn Ser Gly Ala Ser Ser lie Ser 145 150 155 160

Tyr Pro Ala Arg Tyr Ala Asn Ala Met Ala Val Gly Ala Thr Asp Gin 165 170 175

Asn Asn Asn Arg Ala Ser Phe Ser Gin Tyr Gly Ala Gly Leu Asp lie 180 185 190

Val Ala Pro Gly Val Asn Val Gin Ser Thr Tyr Pro Gly Ser Thr Tyr 195 200 205

Ala Ser Leu Asn Gly Thr Ser Met Ala Thr Pro His Val Ala Gly Ala 210 215 220

Ala Ala Leu Val Lys Gin Lys Asn Pro Ser Trp Ser Asn Val Gin He 225 230 235 240

Arg Asn His Leu Lys Asn Thr Ala Thr Ser Leu Gly Ser Thr Asn Leu 245 250 255

Tyr Gly Ser Gly Leu Val Asn Ala Glu Ala Ala Thr Arg 260 265 <21G> 4 <211> 269

<212> PRT <213> Bacillus lentus <400> 4

Ala Gin Ser Val Pro Trp Gly He Ser Arg Val Gin Ala Pro Ala Ala 15 10 15

His Asn Arg Gly Leu Thr Gly Ser Gly Val Lys Val Ala Val Leu Asp 20 25 30

Thr Gly lie Ser Thr His Pro Asp Leu Asn He Arg Gly Gly Ala Ser 35 40 45

Phe Val Pro Gly Glu Pro Ser Thr Gin Asp Gly Asn Gly His Sly Thr 50 SS 60

His Val Ala Gly Thr lie Ala Ala leu Asn Asn Ser lie Sly Val Leu 65 TO 75 80

Gly Val Ala Pro Ser Ala Glu Leu Tyr Ala Val Ays Val leu Sly Ala 85 90 95

Asp Gly Asn Gly Ala lie Ser Ser lie Ala Gin Gly Leu Glu Trp Ala 100 105 110

Gly Asn Asn Gly Met His Val Ala Asn Leu Ser Leu Gly Ser Pro Ser 115 120 125

Pro Ser Ala Tbr leu Glu Gib Ala Val Asn Ser Ala Thr Ser Arg Gly 130 135 140

Val leu Val Val Ala Ala Ser Gly Asn Ser Gly Ala Ser Ser lie Ser 145 150 155 160

Tyr Pro Ala Arg Tyr Ala Asn Ala Met Ala Val Gly Ala Thr Asp Gin 165 170 175

Asn Asn Asn Arg Ala Ser Phe Ser Gin Tyr Gly Ala Gly Leu Asp lie 180 185 190

Val Ala Pro Gly Val Asn val Gin Ser Tbr Tyr Pro Gly Ser Tbr Tyr 195 200 205

Ala Ser Leu Asn Gly Thr Ser Met Ala Thr Pro His Val Ala Gly Ala 210 215 220

Ala Ala Leu Val Lys Gin Lys Asn Pro Ser Trp Ser Asn Val Gift He 225 230 235 240

Arg Asn His Leu Lys Asn Thr Ala Thr Ser Leu Gly Ser Tbr Asn Leu 245 250 255

Tyr Gly Ser Gly Leu Val Asn Ala Glu Ala Ala Tbr Arg 260 265 <210> 5 <211» 269

<212> PRT «213» Bacillus lentus <400» 5

Ala Gin Ser Val Pro Trp Gly lie Ser Arg Val Gin Ala Pro Ala Ala 1 5 10 is

His Asn Arg Gly Leu Thr Gly Sar Gly Val Lys Val Ala Val Lau Asp 20 25 30

Thr Gly Ile Ser Thr His Pro Asp Leu Asn Ile Arg Gly Gly Ala Ser 35 40 45

Phe Val Pro Gly Glu Pro Ser Thr Gin Asp Gly Asn Gly His Gly Thr 50 55 60

His Val Ala Gly Thr Ile Ala Ala Leu Asn Asn Ser Ile Gly Val Leu 65 70 75 80

Gly Val Ala Pro Ser Ala Glu Lau Tyr Ala Val Lys Val Leu Gly Ala 85 50 95

Asp Gly Gin Gly Ala Ile Ser Ser Ile Ala Gin Gly Leu Glu Trp Ala 100 105 110

Gly Asn Asn Gly Met His Val Ala Asn Leu Ser Leu Gly Ser Pro Ser 115 120 125

Pro Ser Ala Thr Leu Glu Gin Ala Val Asn Ser Ala Thr Ser Arg Gly 130 135 140

Val Leu Val Val Ala Ala Ser Gly Asn Ser Gly Ala Ser Ser Ile Ser

145 150 155 ISO

Tyr Pro Ala Arg Tyr Ala Asn Ala Met Ala Val Gly Ala Thr Asp Gin 165 170 175

Asn Asn Asn Arg Ala Ser Phe Ser Gin Tyr Gly Ala Gly Leu Asp Ile 180 185 190

Val Ala Pro Gly Val Asn Val Gin Ser Thr Tyr Pro Gly Ser Thr Tyr 195 200 205

Ala Ser Leu Asn Gly Thr Ser Met Ala Thr Pro His Val Ala Gly Ala 210 215 220

Ala Ala Leu Val Lys Gin Lys Asn Pro Ser Trp Ser Asn Val Gin Ile 225 230 235 240

Arg Asn His Leu Lys Asn Thr Ala Thr Ser Leu Gly Ser Thr Asn Leu 245 250 255

Tyr Gly Ser Gly Leu Val Asn Ala Glu Ala Ala Thr Arg 260 265 <21G> 6 <211> 269

<212> FRT <213> Bacillus lentus <4GQ> 6

Ala Gin Ser Val Pro Trp Gly Ile Ser Arg Val Gin Ala Pro Ala Ala 1 5 10 15

His Asn Arg Gly Leu Thr Gly Ser Gly Val Lys Val Ala Val Leu Asp 20 25 30

Thr Gly Ile Ser Thr His Pro Asp Leu Asn Ile Arg Gly Gly Ala Ser 35 40 45

Phe Val Pro Gly Glu Pro Ser Thr Gin Asp Gly Asn Gly His Gly Thr 50 55 60

His Val Ala Gly Thr lie Ala Ala Leu Asn Asn Ser lie Gly Val Leu 65 70 75 80

Gly Val Ala Pro Ser Ala Glu Leu Tyr Ala Val Lys Val Leu Gly Ala 85 90 95

Asp Gly Ala Gly Ala lie Ser Ser Ik Ala Gin Gly Leu Glu Trp Ala 100 105 110

Gly Asn Asn Gly Met His Val Ala Asn Leu Ser Leu Gly Ser Pro Ser 115 120 125

Pro Ser Ala Thr Leu Glu Gin Ala Val Asn Ser Ala Thr Ser Arg Gly 130 135 140

Val Leu Val Val Ala Ala Ser Gly Asn Ser Gly Ala Ser Ser lie Ser 145 150 155 160

Tyr Pro Ala Arg Tyr Ala Asn Ala Met Ala Val Gly Ala Thr Asp Gin 165 170 175

Asn Asn Asn Arg Ala Ser Phe Ser Gin Tyr Gly Ala Gly Leu Asp lie 180 185 190

Val Ala Pro Gly Val Asn Val Gin Ser Thr Tyr Pro Gly Ser Thr Tyr 195 200 205

Ala Ser Leu Asn Gly Thr Ser Met Ala Thr Pro His Val Ala Gly Ala 210 215 220

Ala Ala Leu Val Lys Gin Lys Asn Pro Ser Trp Ser Asn Val Gin lie 225 230 235 240

Arg Asn His Leu Lys Asn Thr Ala Thr Ser Leu Gly Ser Thr Asn Leu 245 250 255

Tyr Gly Ser Gly Leu Val Asn Ala Glu Ala Ala Thr Arg 260 265 <21G> 7 <211> 269

<212> PRT <213> Bacillus lentus <400> 7

Ala Gin Ser Val Pro Trp Gly Ile Ser Arg Val Gin Ala Pro Ala Ala 15 10 15

His Asn Arg Gly I>eu Thr Gly Ser Gly Val Lys Val Ala Val Leu Asp 20 25 30

Thr Gly Ile Ser Thr His Pro Asp leu Asn Ile Arg Gly Gly Ala Ser 35 40 45

Phe Val Pro Gly Glu Pro Ser Thr Gin Asp Gly Asn Gly His Gly Thr 50 55 60

His Val Ala Gly Thr Ile Ala Ala Le« Asn Asn Ser Ile Gly Val Leu 65 70 75 80

Gly Val Ala Pro Ser Ala Glu Le« Tyr Ala Val Lys Val Leu Gly Ala 85 90 95

Asp Gly Gly Gly Ala Ile Ser Ser Ile Ala Gin Gly Le« Glu Trp Ala 100 105 110

Gly Asn Asn Gly Met His Val Ala Asn Leu Ser Leu Gly Ser Pro Ser 115 120 125

Pro Ser Ala Thr Leu Glu Gin Ala Val Asn Ser Ala Thr Ser Arg Gly 130 135 140

Val Leu Val Val Ala Ala Ser Gly Asn Ser Gly Ala Ser Ser Ile Ser 145 150 155 160

Tyr Pro Ala Arg Tyr Ala Asn Ala Met Ala Val Gly Ala Thr Asp Gin 165 170 175

Asn Asn Asn Arg Ala Ser Phe Ser Gin Tyr Gly Ala Gly Le« Asp Ile 180 185 190

Val Ala Fro Sly Val Asn Val Gin Ser Thr Tyr Fro Gly Ser Thr Tyr 195 200 205

Ala Ser Leu Asn Gly Thr Ser Mat. Ala Thr Fro His Val Ala Gly Ala 210 215 220

Ala Ala Leu Val Lys Gin Lys Asn Fro Ser Trp Ser Asn Val Gin Ile 225 230 235 240

Arg Asn His Len Lys Asn Thr Ala Thr Ser Len Gly Ser Thr Asn Leu 245 250 255

Tyr Gly Ser Gly Leu Val Asn Ala Glu Als Ala Thr Arg 260 265 <21G> 8 <211> 269

<212> PRT <213> Bacillus lentus <40G> 8

Ala Gin Ser Val Pro Trp Gly Ile Ser Arg Val Gin Ala Pro Ala Ala 15 10 15

His Asn Arg Gly Leu Thr Gly Ser Gly Val Lys Val Ala Val Len Asp 20 25 30

Thr Gly Ile Ser Thr His Fro Asp Leu Asn Ile Arg Gly Gly Ala Ser 35 40 45

Phe Val Pro Gly Glu Pro Ser Thr Gin Asp Gly Asn Gly His Gly Thr 50 55 60

His Val Ala Gly Thr Ile Ala Ala Leu Asn Asn Ser Ile Gly Val Leu 65 70 75 80

Gly Val Ala Fro Ser Ala Glu Leu Tyr Ala Val Lys Val Leu Gly Ala 85 90 95

Asp Gly Ser Gly Ala Ile Ser Ser Ile Ala Gin Gly Leu Glu Trp Ala 100 105 110

Gly Asn Asn Gly Met His Val Ala Asn Leu Ser Leu Gly Ser Pro Ser 115 120 125

Pro Ser Ala Thr Leu Glu Gin Ala Val Asn Ser Ala Thr Ser Arg Gly 130 135 140

Val Leu Val Val Ala Ala Ser Gly Asn Ser Gly Ala Ser Ser Ile Ser 145 150 155 160

Tyr Pro Ala Arg Tyr Ala Asn Ala Met Ala Val Gly Ala Thr Asp Gin 165 170 175

Asn Asn Asn Arg Ala Ser Phe Ser Gin Tyr Gly Ala Gly Leu Asp lie ISO 185 190

Val Ala Pro Gly Val Asn Val Gin Ser Thr Tyr Pro Gly Ser Thr Tyr 195 200 205

Ala Ser Leu Asn Gly Thr Ser Met Ala Thr Pro His Val Ala Gly Ala 210 215 220

Ala Ala Leu Val Lys Gin Lys Asn Pro Ser Trp Ser Asn Val Gin He 225 230 235 240

Arg Asn His Leu Lys Asn Thr Ala Thr Ser Leu Gly Ser Thr Asn Leu 245 250 255

Tyr Gly Ser Gly Leu Val Asn Ala Glu Ala Ala Thr Arg 260 265

Claims

Use of a protease (α1) comprising an amino acid sequence at least 80% identical to the amino acid sequence given in SEQ ID NO. 1. and as in position 99 of the count according to SEQ ID NO. 1, the amino acid has glutamic acid (E) or aspartic acid (D), to provide a proteolytic activity in a liquid wash or purification composition, further comprising a phosphonate in an amount of 0.01 to 4% by weight.

The use of claim 1, wherein the protease additionally has at least one of the following amino acids in the count of SEQ ID NO. 1: (a) threonine at position 3 (3T) (b) isoleucine at position 4 (41) (c) alanine, threonine or arginine at position 61 (61A, 61T or 61R) (d) aspartic acid or glutamic acid at position 154 ( 154D or 154E) (e) proline at position 188 (188P) (f) methionine at position 193 (193M) (g) isoleucine at position 199 (1991) (h) aspartic acid, glutamic acid or glycine at position 211 (211D, 211E or 211G) (i) combinations of (a) to (h).

Use of a protease selected from the group consisting of: a. Protease comprising an amino acid sequence of SEQ ID NO. 2 or SEQ ID NO. 3 b. Protease comprising an amino acid sequence modified in at least one position in comparison with SEQ ID NO. 2 or SEQ ID NO. 3, wherein the modification is selected in the count of SEQ ID NO. 1 from the group consisting of: i. Threonine at position 3 (3T) ii. isoleucine at position 4 (41) iii. alanine, threonine or arginine at position 61 (61A, 61T or 61R) iv. aspartic acid or glutamic acid at position 154 (154D or 154E) v. proline at position 188 (188P) vi. methionine at position 193 (193M) vii. isoleucine at position 199 (1991) viii. aspartic acid, glutamic acid or glycine at position 211 (211D, 211E or 211G) ix. combinations of (i) to (viii) to provide a proteolytic activity in a liquid washing or purification composition further comprising a phosphonate in an amount of 0.01 to 4% by weight.

Use of a washing or purifying composition comprising a protease and a phosphonate as claimed in any one of claims 1 to 3, wherein the phosphonate is selected from the group consisting of 1-hydroxyethane-1,1-diphosphonic acid (HEDP), amino tri (methylene phosphonic acid) (ATMP), diethylenetriamine penta (methylene phosphonic acid) (DTPMP or DETPMP or DTPNT), ethylenediamine tetra (methylene phosphonic acid) (EDTMP), 2-phosphonobutane-1,2,4-tricarboxylic acid (PBS) thereof.

Use according to any one of claims 1 to 4, wherein the protease is present in an amount of 1 x 10-8 to 5% by weight based on active protein.

Use according to any one of claims 1 to 5, further comprising a component selected from: i. Anionic and / or polyanionic substance and / or ii. cationic and / or polycationic substance and / or iii. substance with hydroxyl and / or polyhydroxyl group (s).

Use according to any one of claims 1 to 6, comprising at least one additional component, in particular one selected from the group consisting of surfactant, builder, peroxygen compound, bleach activator, alcohol, acid, gray staining inhibitor, optical bleach, foam inhibitor, water-soluble salt, thickener, volatile alkali and / or base, hydrophilizing agent and combinations thereof.

Use according to any of claims 1 to 7, wherein it comprises at least one additional enzyme, in particular a protease, amylase, cellulase, hemicellulase, mannanase, tannase, xylanase, xanathanase, β-glucosidase, carrageenase, perhydrolase, oxidase, oxidoreductase or a lipase and preferably mixtures thereof.

Use according to any one of claims 1 to 8 for the removal of protease-sensitive dirt on textiles or hard surfaces.

A method of cleaning textiles or hard surfaces, wherein a washing or cleaning composition as defined in any one of claims 1 to 8 is used in at least one process step.

The method of claim 10, wherein the phosphonate is present in the wash liquid at a concentration of 0.00075 to 0.05% by weight and / or the protease is present in the wash liquid at a concentration of 0.0005 to 0.03% by weight.

The method of claim 10 or 11, wherein it is carried out at a temperature of between 10 ° C and 60 ° C.